Methods of treating leukocytosis, endothelial dysfunction and carditis using lipid-binding protein-based complexes

Abstract

A method for treating leukocytosis, endothelial dysfunction, and carditis comprises administering to a subject a lipid binding protein based complex.

Description

[0001] 1. Cross-reference to related applications

[0002] This application claims priority to U.S. Provisional Application No. 63 / 328,210, filed on April 6, 2022, the contents of which are incorporated herein by reference in their entirety.

[0003] 2. Sequence Listing

[0004] This application contains a sequence listing submitted electronically, which is incorporated herein by reference in its entirety. The electronic copy was created on April 3, 2023, is an XML file named CRN-048WO_ST26, and is 3,244 bytes in size. 3. Summary of the Invention

[0006] The present disclosure is based in part on the discovery that subjects treated with the lipid binding protein based complex CER-001 as described in Example 2 showed unexpectedly rapid improvements in biomarkers of inflammation, leukocytosis, and endothelial dysfunction.

[0007] Thus, in some aspects, the present disclosure provides methods of treating a subject having or at risk of developing leukocytosis with a dose (eg, a high dose) of a lipid binding protein-based complex.

[0008] In some aspects, the present disclosure provides methods of treating a subject having one or more symptoms associated with leukocytosis, the method comprising administering to the subject a dose of a lipid binding protein-based complex and a dose (e.g., a high dose) of a lipid binding protein-based complex.

[0009] In some aspects, the disclosure provides methods of treating a subject having endothelial dysfunction (e.g., a subject having or having experienced an acute coronary syndrome or stroke) with a dose (e.g., a high dose) of a lipid binding protein-based complex.

[0010] In some aspects, the present disclosure provides methods of treating a subject who is experiencing or has experienced an acute coronary syndrome or stroke with a high dose of a lipid binding protein-based complex.

[0011] In some aspects, the present disclosure provides methods of treating a subject having or at risk of developing carditis with a dose (eg, a high dose) of a lipid binding protein-based complex.

[0012] In the method of the present disclosure, the lipid-binding protein-based complex is usually administered in high doses. High doses are usually higher than the doses used to treat chronic conditions such as familial hypercholesterolemia. The high dose is usually administered in a relatively short period of time, for example, in a period of two days to two weeks, and usually includes multiple administrations of lipid-binding protein-based complexes, for example, two to 20 separate doses. Each dose can be spaced less than one day (e.g., twice a day) or one day or longer (e.g., once a day).

[0013] In some embodiments of the methods of the present disclosure, the lipid binding protein-based complex comprises sphingomyelin and / or a negatively charged lipid, such as CER-001. CER-001 is a negatively charged lipoprotein complex comprising recombinant human ApoA-I, sphingomyelin (SM) and 1,2-dihexadecanoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (dipalmitoylphosphatidyl-glycerol; DPPG). It mimics native, nascent discoid pre-βHDL, which is the form that HDL particles take before acquiring cholesterol. Without being bound by theory, it is believed that CER-001 therapy can (1) reduce serum levels of inflammatory cytokines such as IL-6, (2) reduce white blood cell counts in subjects with or at risk of leukocytosis, and (3) reduce serum levels of ICAM-1 and VCAM-1, thereby providing clinical benefits to subjects with or at risk of developing leukocytosis, subjects with endothelial dysfunction, and subjects with or at risk of developing carditis.

[0014] In some aspects, the disclosure provides dosing regimens for a lipid binding protein-based therapy (eg, CER-001 therapy) in a subject as described herein.

[0015] The dosing regimen of the present disclosure generally requires multiple administrations of CER-001 to the subject (e.g., daily administration or administration about 12 hours apart). CER-001 therapy can continue for a predetermined period of time, such as a week or a period of time exceeding a week (e.g., two weeks). Alternatively, CER-001 can be continuously administered to the subject until one or more symptoms of the condition experienced by the subject are alleviated or continued until the serum level of one or more inflammatory markers is reduced, such as to normal levels or relative to a baseline measurement taken before the start of CER-001 treatment. For subjects with infection, in some embodiments, therapy can be continued until the subject recovers from the infection.

[0016] Dosing regimens of the present disclosure may entail administering the lipid binding protein-based complex (eg, CER-001) to the subject according to an initial "induction" regimen, optionally followed by administering the lipid binding protein-based complex to the subject according to a "consolidation" regimen.

[0017] The induction regimen typically includes administering multiple doses of the lipid binding protein-based complex (eg, CER-001) to the subject, for example, six doses over three days.

[0018] The consolidation regimen generally includes administering one or more doses of a lipid-binding protein-based complex (e.g., CER-001) to the subject after the final dose of the induction regimen, for example, one or more days after the final dose of the 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 lipid-binding protein-based complex (e.g., CER-001) to the subject according to the induction regimen on days 1, 2, and 3, and administering a lipid-binding protein-based complex to the subject according to the consolidation regimen on day 6. In some embodiments, the consolidation regimen includes two doses of a lipid-binding protein-based complex.

[0019] In certain embodiments, the present disclosure provides methods of treating a subject as described herein with a lipid binding protein-based complex (e.g., CER-001) according to a dosage regimen comprising:

[0020] - 2 doses per day on days 1, 2 and 3 (induction regimen), optionally followed by

[0021] - 2 subsequent doses on Day 4 or later (consolidation regimen).

[0022] In some embodiments, the regimen comprises:

[0023] -2 doses per day on days 1, 2, and 3 (induction regimen), followed by

[0024] - 2 doses on Day 6 (consolidation regimen).

[0025] In some embodiments, the dosing regimen comprises a single phase, eg, corresponding to an induction regimen described herein.

[0026] In certain aspects, the lipid binding protein-based complex (eg, CER-001) is administered in combination with standard of care therapy (eg, antibiotic therapy for infection and / or hemodynamic support).

[0027] In certain aspects, an antihistamine (eg, dexchlorpheniramine, hydroxyzine, diphenhydramine, cetirizine, fexofenadine, or loratadine) can be administered prior to administration of the lipid binding protein-based complex (eg, CER-001). Antihistamines can reduce the likelihood of an allergic reaction. 4. Description of the drawings

[0028] Figure 1: Showing IL-6 serum levels in a porcine model of sepsis-induced AKI (Example 1).

[0029] Figure 2 : Showing soluble VCAM-1 serum levels in a porcine model of sepsis-induced AKI (Example 2).

[0030] Figure 3 : Showing soluble ICAM-1 serum levels in a porcine model of sepsis-induced AKI (Example 3).

[0031] Figure 4 : Showing LPS serum levels in a porcine model of sepsis-induced AKI (Example 1).

[0032] Figure 5 : A schematic diagram showing the clinical study of Example 2 is shown.

[0033] Figure 6 This is a flow chart used for the study of Example 3.

[0034] Figure 7 This is a flow chart used for the study of Example 4.

[0035] Figures 8A-8J The results show that MCP1 ( Fig. 8A )、TNF-α( Figure 8B )、VCAM( Figure 8C )、ICAM( Fig.8D )、Ferritin( Fig. 8E ),leukocyte( Figure 8F )、CRP( Figure 8G )、KIM-1( Figure 8H )、IL-8( Figure 8I ) and triglycerides ( Figure 8J ) changes.

[0036] Figures 9A-9F The changes in VCAM in the standard of care (SOC) group and the three CER-001 groups in the clinical study of Example 2 are shown. Fig.9A , change from baseline in the SOC group compared with the combined CER-001 group. Fig. 9B , changes from baseline in each group. Fig. 9C , percentage changes in peak values ​​for the SOC group and the combined study groups. Fig.9D , the percentage change in the peak value between the SOC group and the study group, according to whether the subjects were admitted from the ICU or the nephrology department of the center. Fig.9E , SOC, and the change from baseline for each subject in the combined CER-001 groups. Fig.9F Change from baseline for each subject in the SOC group and in each study group.

[0037] Figures 10A-10F The changes in ICAM in the standard of care (SOC) group and the three CER-001 groups in the clinical study of Example 2 are shown. Fig. 10A , change from baseline in the SOC group compared with the combined CER-001 group. Fig. 10B , changes from baseline in each group. Fig. 10C , percentage changes in peak values ​​for the SOC group and the combined study groups. Fig. 10D , the percentage change in the peak value between the SOC group and the study group, according to whether the subjects were admitted from the ICU or the nephrology department of the center. Fig.10E , SOC, and the change from baseline for each subject in the combined CER-001 groups. Fig.10F Change from baseline for each subject in the SOC group and in each study group.

[0038] Fig.11 Shown are the results of an MTT cell viability assay of cultured endothelial cells following challenge with LPS and CER-001 infusion, as described in Example 6.

[0039] Fig.12 The results of endothelial nitric oxide synthase (eNOS)-based FACS (eNOS (phospho S1177)) of cultured endothelial cells after challenge with LPS and CER-001 infusion, as described in Example 6, are summarized.

[0040] Fig.13 Shown are the results of eNOS (phosphorylated S1177) based FACS of cultured endothelial cells challenged with LPS and CER-001 infusion as described in Example 6, compared to basal cells and VEFG (positive control) cells in one representative experiment of three independent experiments.

[0041] Fig.14 Shown are the results of an MTT cell viability assay of PBMCs from healthy donors following challenge with LPS and CER-001 infusion, as described in Example 6.

[0042] Fig.15 TNF-α synthesis by PBMCs from healthy donors after challenge with LPS and CER-001 infusion, as described in Example 6, is shown.

[0043] Fig.16 Shown are the results of CD14-based FACS of PBMCs from healthy donors following challenge with LPS and CER-001 infusion, as described in Example 6, in one representative experiment of three independent experiments.

[0044] Fig.17The results of CD14-based FACS of PBMCs from healthy donors after challenge with LPS and CER-001 infusion, as described in Example 6, are summarized.

[0045] Figures 18A-18F The changes in white blood cell counts of the standard care (SOC) group and the CER-001 group in the clinical study of Example 2 are shown. Fig.18A : Changes from baseline in white blood cell counts in the SOC group and the combined CER-001 group. Fig.18B : Change from baseline in white blood cell counts in the SOC group and each CER-001 group. Fig.18C : Change in white blood cell count for SOC and combined CER-001 groups, reported as a percentage of peak white blood cell count (peak = 100%). Treatment x study day effect relative to peak p = 0.5492. Fig.18D : Changes in white blood cell counts in the SOC group and the combined CER-001 group, according to whether the subjects were enrolled from the ICU or the nephrology department of the center. Fig.18E : Fig.18A The individual data points are summarized in . Fig.18F : Fig.18B The individual data points are summarized in . 5. Detailed description of the invention

[0047] In some aspects, the present disclosure provides methods of treating a subject having or at risk of developing leukocytosis with a dose (eg, a high dose) of a lipid binding protein-based complex.

[0048] In some aspects, the present disclosure provides methods for treating a subject having one or more symptoms associated with leukocytosis, the method comprising administering to the subject a dose of a lipid-binding protein-based complex and a dose (e.g., a high dose) of a lipid-binding protein-based complex.

[0049] In some aspects, the disclosure provides methods of treating a subject having endothelial dysfunction (e.g., a subject having or having experienced an acute coronary syndrome or stroke) with a dose (e.g., a high dose) of a lipid binding protein-based complex.

[0050] In some aspects, the present disclosure provides methods of treating a subject who is experiencing or has experienced an acute coronary syndrome or stroke with a high dose of a lipid binding protein-based complex.

[0051] In some aspects, the present disclosure provides methods of treating a subject having or at risk of developing carditis with a dose (eg, a high dose) of a lipid binding protein-based complex.

[0052] In some embodiments of the methods of the present disclosure, the lipid binding protein-based complex is an apomer, a cargomer, a HDL-based complex, or a HDL mimetic-based complex. In specific embodiments, the lipid binding protein-based complex is CER-001.

[0053] Exemplary features of lipid binding protein-based complexes useful in the methods and compositions of the present disclosure are described in Section 5.1. Exemplary subject populations that can be treated by the methods of the present disclosure and with the compositions of the present disclosure are described in Section 5.2.

[0054] In some embodiments, the methods of the present disclosure include administering a lipid-binding protein-based complex (e.g., CER-001) to a subject in two stages. First, a lipid-binding protein-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, a lipid-binding protein-based complex (e.g., CER-001) can be administered to a subject in a single stage, for example, according to an administration regimen corresponding to the dose and frequency of administration of the induction or consolidation regimen described herein.

[0055] Induction regimens that can be used in the methods of the present disclosure are described in Section 5.3, and consolidation regimens that can be used in the methods of the present disclosure are described in Section 5.3.2. Dosing regimens of the present disclosure include administration of lipid-binding protein-based complexes (e.g., CER-001) as a monotherapy or as part of a combination therapy with one or more drugs, such as in combination with standard of care therapy for sepsis or other infections (e.g., antibiotic therapy and / or hemodynamic support). Combination therapy is described in Section 5.4.

[0056] 5.1. Lipid-binding protein-based complexes

[0057] 5.1.1. Composites based on HDL and HDL mimetics

[0058] In one aspect, lipid-binding protein-based complexes include complexes based on HDL or HDL mimetics. For example, the complex may include lipoprotein complexes as described in US Patent No. 8,206,750, PCT Publication WO 2012 / 109162, PCT Publication WO 2015 / 173633A2 (e.g., CER-001), or US 2004 / 0229794A1, each of which is incorporated herein by reference as a whole. The terms "lipoprotein" and "apolipoprotein" are used interchangeably herein, and unless the context otherwise requires, the term "lipoprotein" covers lipoprotein mimetics. The terms "lipid-binding protein" and "lipid-binding polypeptide" are also used interchangeably herein, and unless the context otherwise requires, these terms do not mean amino acid sequences of a specific length.

[0059] The lipoprotein complex may comprise a protein fraction (e.g., an apolipoprotein fraction) and a lipid fraction (e.g., a phospholipid fraction). The protein fraction includes one or more lipid-binding protein molecules, such as apolipoprotein, a peptide, or an apolipoprotein peptide analog or mimetic, such as one or more lipid-binding protein molecules described in Section 5.1.2.

[0060] The lipid fraction typically includes one or more phospholipids, which may be neutral, negatively charged, positively charged, or a combination thereof. Exemplary phospholipids and other amphiphilic molecules that may be included in the lipid fraction are described in Section 5.1.4.

[0061] In certain embodiments, the lipid fraction comprises at least one neutral phospholipid (e.g., sphingomyelin (SM)) and optionally one or more negatively charged phospholipids. In lipoprotein complexes including neutral and negatively charged phospholipids, the neutral and negatively charged phospholipids can have fatty acid chains with the same or different carbon numbers and the same or different saturation. In some cases, the neutral and negatively charged phospholipids will have the same acyl tail, such as C16:0 or palmitoyl, acyl chain. In specific embodiments, particularly those in which SM is used as a neutral lipid, the weight ratio of the apolipoprotein fraction: lipid fraction is about 1:2.7 to about 1:3 (e.g., 1:2.7).

[0062] Any phospholipid with at least a partial negative charge at physiological pH can be used as a negatively charged phospholipid. Non-limiting examples include negatively charged forms of phosphatidylinositol, phosphatidylserine, phosphatidylglycerol, and phosphatidic acid, such as salts. In a specific embodiment, the negatively charged phospholipid is 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] or DPPG (a phosphatidylglycerol). Preferred salts include potassium salts and sodium salts.

[0063] In some embodiments, the lipoprotein complex used in the methods of the present disclosure is a lipoprotein complex as described in US Patent No. 8,206,750 or WO 2012 / 109162 (and its US counterpart, US2012 / 0232005), the contents of each of which are incorporated herein by reference in their entirety. In a specific embodiment, the protein component of the lipoprotein complex is as described in Section 6.1 of WO 2012 / 109162 (and US2012 / 0232005) and preferably as described in Section 6.1.1, and the lipid component is as described in Section 6.2 of WO 2012 / 109162 (and US2012 / 0232005), which may optionally be compounded together in the amounts described in Section 6.3 of WO 2012 / 109162 (and US 2012 / 0232005). The contents of each of these sections are 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 contents of which are incorporated herein by reference.

[0064] 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 phosphatidylcholine molecules, and 20-50 SM molecules.

[0065] 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 phosphatidylcholine molecules, and 50 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, 80 phosphatidylcholine molecules, and 20 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, 70 phosphatidylcholine molecules, and 30 SM molecules.

[0068] 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 phosphatidylcholine molecules, and 40 SM molecules.

[0069] In a specific embodiment, the lipoprotein complex useful in the methods of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 50-80 phosphatidylcholine molecules, and 20-50 SM molecules.

[0070] In another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 50 phosphatidylcholine molecules, and 50 SM molecules.

[0071] In yet another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 80 phosphatidylcholine molecules, and 20 SM molecules.

[0072] In yet another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 70 phosphatidylcholine molecules, and 30 SM molecules.

[0073] In yet another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 60 phosphatidylcholine molecules, and 40 SM molecules.

[0074] In a specific embodiment, the lipoprotein complexes useful in the methods of the present disclosure comprise a lipid component comprising about 90 wt% to 99.8 wt% SM and about 0.2 wt% to 10 wt% negatively charged phospholipids, for example, about 0.2 wt%-1 wt%, 0.2 wt%-2 wt%, 0.2 wt%-3 wt%, 0.2 wt%-4 wt%, 0.2 wt%-5 wt%, 0.2 wt%-6 wt%, 0.2 wt%-7 wt%, 0.2 wt%-8 wt%, 0.2 wt%-9 wt% or 0.2 wt%-10 wt% total negatively charged phospholipids. In another specific embodiment, the lipoprotein complexes useful in the methods of the present disclosure contain about 90 wt% to 99.8 wt% phosphatidylcholine and about 0.2 wt% to 10 wt% negatively charged phospholipids, for example, about 0.2 wt%-1 wt%, 0.2 wt%-2 wt%, 0.2 wt%-3 wt%, 0.2 wt%-4 wt%, 0.2 wt%-5 wt%, 0.2 wt%-6 wt%, 0.2 wt%-7 wt%, 0.2 wt%-8 wt%, 0.2 wt%-9 wt% or 0.2 wt%-10 wt% total negatively charged phospholipids.

[0075] In a specific embodiment, the lipoprotein complexes useful in the methods of the present disclosure comprise a lipid component consisting essentially of about 90 wt% to 99.8 wt% SM and about 0.2 wt% to 10 wt% negatively charged phospholipids, for example, about 0.2 wt%-1 wt%, 0.2 wt%-2 wt%, 0.2 wt%-3 wt%, 0.2 wt%-4 wt%, 0.2 wt%-5 wt%, 0.2 wt%-6 wt%, 0.2 wt%-7 wt%, 0.2 wt%-8 wt%, 0.2 wt%-9 wt% or 0.2 wt%-10 wt% total negatively charged phospholipids. In another specific embodiment, the lipoprotein complexes useful in the methods of the present disclosure consist essentially of about 90 wt% to 99.8 wt% phosphatidylcholine and about 0.2 wt% to 10 wt% negatively charged phospholipids, for example, about 0.2 wt%-1 wt%, 0.2 wt%-2 wt%, 0.2 wt%-3 wt%, 0.2 wt%-4 wt%, 0.2 wt%-5 wt%, 0.2 wt%-6 wt%, 0.2 wt%-7 wt%, 0.2 wt%-8 wt%, 0.2 wt%-9 wt% or 0.2 wt%-10 wt% total negatively charged phospholipids.

[0076] In yet another specific embodiment, the lipoprotein complexes useful in the methods of the present disclosure comprise a lipid fraction comprising about 9.8 wt% to 90 wt% SM, about 9.8 wt% to 90 wt% phosphatidylcholine, and about 0.2 wt% to 10 wt% negatively charged phospholipids, for example, about 0.2 wt%-1 wt%, 0.2 wt%-2 wt%, 0.2 wt%-3 wt%, 0.2 wt%-4 wt%, 0.2 wt%-5 wt%, 0.2 wt%-6 wt%, 0.2 wt%-7 wt%, 0.2 wt%-8 wt%, 0.2 wt%-9 wt%, to 0.2 wt%-10 wt% total negatively charged phospholipids.

[0077] In yet another specific embodiment, the lipoprotein complexes useful in the methods of the present disclosure comprise a lipid fraction consisting essentially of about 9.8 wt% to 90 wt% SM, about 9.8 wt% to 90 wt% phosphatidylcholine, and about 0.2 wt% to 10 wt% negatively charged phospholipids, for example, about 0.2 wt%-1 wt%, 0.2 wt%-2 wt%, 0.2 wt%-3 wt%, 0.2 wt%-4 wt%, 0.2 wt%-5 wt%, 0.2 wt%-6 wt%, 0.2 wt%-7 wt%, 0.2 wt%-8 wt%, 0.2 wt%-9 wt%, to 0.2 wt%-10 wt% negatively charged phospholipids.

[0078] In another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure comprises ApoA-I apolipoprotein and a lipid fraction, wherein the lipid fraction comprises 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 disc-shaped particle containing 2-4 ApoA-I equivalents.

[0079] In another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure comprises ApoA-I apolipoprotein and a lipid fraction, wherein the lipid fraction consists essentially of 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 disc-shaped particle containing 2-4 ApoA-I equivalents.

[0080] HDL-based or HDL-mimic-based complexes may include a single type of lipid-binding protein, or a mixture of two or more different lipid-binding proteins, which may be derived from the same or different species. Although not required, the complex will preferably contain a lipid-binding protein derived from or corresponding to the animal species being treated in amino acid sequence to avoid inducing an immune response to therapy. Therefore, for the treatment of human patients, lipid-binding proteins of human origin are preferably used. The use of peptide mimics of apolipoproteins can also reduce or avoid immune responses.

[0081] 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 5.1.4.1.

[0082] The lipid component comprising SM may optionally include a small amount of additional lipids. Virtually any type of lipid may be used, including but not limited to lysophospholipids, galactocerebrosides, gangliosides, cerebrosides, glycerides, triglycerides and cholesterol and their derivatives.

[0083] When included, such optional lipids typically comprise less than about 15 wt % of the lipid fraction, although more optional lipids may be included in some cases. In some embodiments, the optional lipids comprise less than about 10 wt %, less than about 5 wt %, or less than about 2 wt %. In some embodiments, the lipid fraction does not include optional lipids.

[0084] In a specific embodiment, the phospholipid fraction comprises egg SM or palmitoyl SM or phytosphingomyelin and DPPG, with a weight ratio (SM: negatively charged phospholipid) ranging from 90:10 to 99:1, more preferably ranging from 95:5 to 98:2. In one embodiment, the weight ratio is 97:3.

[0085] The molar ratio of the lipid component to the protein component of the complex of the present disclosure can vary and will depend on factors such as the properties of the apolipoprotein comprising the protein component, the properties and amount of the lipid comprising the lipid component, and the desired size of the complex. Because the biological activity of apolipoproteins such as ApoA-I is believed to be mediated by the amphipathic helices comprising the apolipoprotein, it is convenient to express the lipid: apolipoprotein portion of the apolipoprotein molar ratio using ApoA-I protein equivalents. ApoA-I is generally believed to contain 6-10 amphipathic helices, depending on the method used to calculate the helices. Other apolipoproteins can be expressed in the form of ApoA-I equivalents based on the number of amphipathic helices they contain. For example, ApoA-I, which generally exists as a disulfide-bridged dimer, is expressed in the form of apolipoprotein equivalents. M It can be expressed as 2 ApoA-I equivalents, because each ApoA-I M The molecule contains twice as many amphipathic helices as an ApoA-I molecule. In contrast, a peptide apolipoprotein containing a single amphipathic helix can be expressed as 1 / 10-1 / 6 ApoA-I equivalents because each molecule contains 1 / 10-1 / 6 of the amphipathic helices of an ApoA-I molecule. Typically, the lipid: ApoA-I equivalent molar ratio (defined herein as "Ri") in the lipoprotein complex will be in the range of about 105:1 to 110:1. In some embodiments, Ri is about 108:1. For phospholipids, a MW of about 650-800 can be used to obtain the weight ratio.

[0086] In some embodiments, the molar ratio of lipid:ApoA-I equivalents ("RSM") ranges from about 80:1 to about 110:1, such as about 80:1 to about 100: 1. In a specific example, the RSM of the complex can be about 82:1.

[0087] In some embodiments, the lipoprotein complex used in the methods of the present disclosure is a negatively charged complex comprising a protein fraction, preferably mature full-length ApoA-I, and a lipid fraction comprising neutral phospholipids, sphingomyelin (SM) and negatively charged phospholipids.

[0088] In a specific embodiment, the lipid component comprises SM (e.g., egg SM, palmitoyl SM, plant SM, or a combination thereof) and a negatively charged phospholipid (e.g., DPPG) in a weight ratio (SM: negatively charged phospholipid) ranging from 90:10 to 99:1, more preferably 95:5 to 98:2, e.g., 97:3.

[0089] In specific embodiments, the ratio of the protein component to the lipid component can be in the range of about 1:2.7 to about 1:3, with 1:2.7 being preferred. This corresponds to a molar ratio of ApoA-I protein to lipid in the range of about 1:90 to 1:140. In some embodiments, the molar ratio of protein to lipid in the complex is about 1:90 to about 1:120, about 1:100 to about 1:140, or about 1:95 to about 1:125.

[0090] In particular embodiments, the complex comprises CER-001, CSL-111, CSL-112, CER-522, or ETC-216. In preferred embodiments, the complex is CER-001.

[0091] CER-001 used in the literature and in the following examples 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 a SM: DPPG weight: weight ratio of 97: 3. Example 4 of WO 2012 / 109162 also describes its production method.

[0092] 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 vary by up to 20% from CER-001 as described in Example 4 of WO 2012 / 109162. In certain embodiments, the components of the lipoprotein complex vary by up to 10% from CER-001 as described in Example 4 of WO 2012 / 109162. Preferably, the components of the lipoprotein complex are those described in Example 4 of WO 2012 / 109162 (plus / minus acceptable manufacturing tolerance variations). The SM in CER-001 may be natural or synthetic. In some embodiments, the SM is a natural SM, such as a natural SM described in WO 2012 / 109162, such as an egg SM. In some embodiments, the SM is a synthetic SM, such as a synthetic SM described in WO 2012 / 109162, such as a synthetic palmitoyl sphingomyelin, for example, as described in WO 2012 / 109162. Methods for synthesizing palmitoyl sphingomyelin are known in the art, for example, as 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:1 of WO 2012 / 109162 (same as SEQ ID NO:2 of the present application). ApoA-I can be purified or recombinantly produced by animal sources (particularly human sources). In a preferred embodiment, the ApoA-I in CER-001 is recombinant ApoA-I. CER-001 used in the dosing regimens of the present disclosure is preferably highly homogeneous, e.g., 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, e.g., Section 6.4 of WO 2012 / 109162.

[0093] CSL-111 is reconstituted human ApoA-I purified from plasma complexed with soybean phosphatidylcholine (SBPC) (Tardif et al., 2007, JAMA 297: 1675-1682).

[0094] 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).

[0095] ETC-216 (also known as MDCO-216) is a recombinant ApoA-I MilanoA lipid-depleted form of HDL. See Nicholls et al., 2011, Expert Opin Biol Ther. 11(3):387-94. doi: 10.1517 / 14712598.2011.557061.

[0096] 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 that contains a combination of three phospholipids and a 22 amino acid peptide CT80522:

[0097]

[0098] The phospholipid component of CER-522 is composed of egg sphingomyelin, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (dipalmitoylphosphatidylcholine, DPPC) and 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (dipalmitoylphosphatidyl-glycerol, 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).

[0099] In some embodiments, the lipoprotein complex is delipidated HDL. Most HDL in plasma is rich in cholesterol. Lipids in HDL can be depleted, such as partially and / or selectively depleted, such as to reduce its cholesterol content. In some embodiments, delipidated HDL can be similar to small alpha, preβ-1 and other preβ forms of HDL. Methods for selectively depleting HDL are described in Sacks et al., 2009, J Lipid Res. 50(5): 894–907.

[0100] In certain embodiments, the lipoprotein complex comprises a bioactive agent delivery particle as described in US 2004 / 0229794.

[0101] The bioactive agent delivery particle can comprise a lipid-binding polypeptide (e.g., an apolipoprotein as described previously in this section or in Section 5.1.2), a lipid bilayer (e.g., comprising one or more phospholipids as described previously in this section or in Section 5.1.4.1), and a bioactive agent (e.g., an anticancer 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 particle is as described in US2004 / 0229794.

[0102] In some embodiments, the bioactive agent delivery particle does not comprise a hydrophilic core.

[0103] In some embodiments, the bioactive agent delivery particles are disc-shaped (eg, having a diameter of about 7 to about 29 nm).

[0104] The bioactive agent delivery particles include bilayer-forming lipids, such as phospholipids (e.g., as previously described in this section or in Section 5.1.4.1). In some embodiments, the bioactive agent delivery particles include bilayer-forming 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 comprises DMPC and DMPG in a molar ratio of 7:3.

[0105] In some embodiments, the lipid-binding polypeptide is an apolipoprotein (e.g., as described previously in this section or in Section 5.1.2). The primary interaction between a lipid-binding polypeptide (e.g., an apolipoprotein molecule) and a lipid bilayer is typically a hydrophobic interaction between residues on the hydrophobic face of an amphipathic structure (e.g., an α-helix of the lipid-binding polypeptide) and the fatty acyl chains of lipids on the outer surface of the particle periphery. The bioactive agent delivery particle may include exchangeable and / or non-exchangeable apolipoproteins. In one embodiment, the lipid-binding polypeptide is ApoA-I.

[0106] In some embodiments, the bioactive agent delivery particle includes a lipid-binding polypeptide molecule, such as an apolipoprotein molecule, that has been modified to increase particle stability. In one embodiment, the modification includes the introduction of cysteine ​​residues to form intramolecular and / or intermolecular disulfide bonds.

[0107] In another embodiment, the bioactive agent delivery particle comprises a chimeric lipid-binding polypeptide molecule, such as a chimeric apolipoprotein molecule, having one or more associated functional moieties, such as one or more targeting moieties and / or one or more moieties having a desired biological activity (e.g., antimicrobial activity), which can enhance the activity of or act synergistically with the bioactive agent incorporated into the delivery particle.

[0108] 5.1.2. ApoA-I preparations

[0109] In one aspect, the disclosure relates to an 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.

[0110] ApoA-I can be any such apolipoprotein described in Section 5.1.3.1, including but not limited to ApoA-I having an amino acid sequence of amino acids 25-267 of SEQ ID NO: 2 and / or recombinantly expressed ApoA-I.

[0111] The lipid can be any one or more of those described in Section 5.1.4.1. The lipid can include a neutral lipid and / or a negatively charged lipid. The neutral lipid can include or consist of a sphingomyelin, such as a natural sphingomyelin (e.g., egg sphingomyelin) and / or a synthetic sphingomyelin (e.g., palmitoyl sphingomyelin). The negatively charged lipid can include 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] ("DPPG") or a salt thereof, or consist of DPPG or a salt thereof.

[0112] Lipid can comprise any weight as described herein or the neutral phospholipid of mol ratio and negatively charged phospholipid.Lipid can be made up of 95 % by weight to 99 % by weight neutral phospholipid and 1 % by weight to 5 % by weight negatively charged phospholipid, for example 96 % by weight to 98 % by weight neutral phospholipid and 2 % by weight to 4 % by weight negatively charged phospholipid, or 97 % by weight neutral phospholipid and 3 % by weight negatively charged phospholipid.

[0113] The formulation may include ApoA-I and lipids in any weight or molar ratio described herein. For example, the molar ratio of the components of negatively charged lipids to neutral lipids to ApoA-I in the formulation is 2-6:90-120:1. Exemplary formulations may include ApoA-I to lipids in a ratio ranging from 1:2 to 1:3 by weight, for example, about 1:2.7 by weight.

[0114] The formulations can be used in methods of treating a subject who has one or more symptoms associated with leukocytosis, has endothelial dysfunction, has carditis or is at risk of developing carditis, has leukocytosis or is at risk of developing leukocytosis, is experiencing an acute coronary syndrome or stroke, and / or has experienced an acute coronary syndrome or stroke, such as the methods described in Sections 5.2-5.4. The formulations can also be used to treat such diseases or conditions. The formulations can also be used to prepare a medicament for treating such diseases or conditions.

[0115] Examples of ApoA-I preparations and uses thereof include those described in Group 2, numbered embodiments 1-72.

[0116] 5.1.3. Lipid binding protein molecules

[0117] Lipid binding protein molecules that can be used in the complexes described herein include those apolipoproteins as described in Section 5.1.3.1 and those apolipoprotein mimetic peptides as described in Section 5.1.3.2. In some embodiments, the complex comprises a mixture of lipid binding protein molecules. In some embodiments, the complex comprises a mixture of one or more lipid binding protein molecules and one or more apolipoprotein mimetic peptides.

[0118] 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). Lipid binding proteins 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 It can be expressed as 2 ApoA-I equivalents, because each ApoA-I M The molecule contains twice as many amphipathic helices as an ApoA-I molecule. In contrast, a peptide mimetic containing a single amphipathic helix can be expressed as 1 / 10-1 / 6 ApoA-I equivalents, since each molecule contains 1 / 10-1 / 6 of the amphipathic helices of an ApoA-I molecule.

[0119] 5.1.3.1. Apolipoprotein

[0120] Suitable apolipoproteins that can be included in the lipid-binding protein-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 above. Polymorphic forms, isotypes, variants and mutants and truncated forms of the aforementioned apolipoproteins can also be used, among which the most common 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). Apolipoprotein can be in the form of a monomer or dimer, which can be a homodimer or a heterodimer. For example, 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., 1985, J. Biol. Chem. 260: 1632-35) can be used. 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., 1985, J. Biol. Chem. 260 (14): 8637-46) al., 1991, Euro. J. Biochem. 201(2):373-83), ApoE (McLean et al., 1983, J. Biol. Chem. 258(14):8993-9000), homo- and heterodimers of ApoJ and ApoH (where feasible).

[0121] Apolipoprotein can be modified in its primary sequence to make it less susceptible to oxidation, for example, as described in US Publication Nos. 2008 / 0234192 and 2013 / 0137628 and US Patent Nos. 8,143,224 and 8,541,236. Apolipoprotein can include residues corresponding to elements that facilitate its separation, such as a His tag, 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)).

[0122] In some embodiments, the complex comprises covalently bound lipid binding protein monomers, such as dimeric apolipoprotein A-IMilano, which is a mutant form of ApoA-I containing cysteine. Cysteine ​​allows the formation of disulfide bridges, which can lead to the formation of homodimers or heterodimers (e.g., ApoA-I Milano-ApoA-II).

[0123] 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.

[0124] In some embodiments, the apolipoprotein molecule comprises 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.

[0125] In some embodiments, the ApoA-I molecule is an apolipoprotein A-IMilano (ApoA-IM), apolipoprotein A-IParis (ApoA-IP), or an apolipoprotein A-IZaragoza (ApoA-IZ) molecule.

[0126] Apolipoproteins can be purified from animal sources (particularly from human sources) or recombinantly produced, as is 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 US Patent Nos. 5,059,528, 5,128,318, 6,617,134; US Publication Nos. 2002 / 0156007, 2004 / 0067873, 2004 / 0077541, and 2004 / 0266660; and PCT Publication Nos. WO 2008 / 104890 and WO 2007 / 023476. Other purification methods are also possible, for example as described in PCT Publication No. WO 2012 / 109162, the disclosure of which is incorporated herein by reference in its entirety.

[0127] Apolipoprotein can be in a pro-form, a precursor form, or a mature form. For example, the complex can include ApoA-I (e.g., human ApoA-I), wherein ApoA-I is pre-pro-ApoA-I, pre-ApoA-I, or mature ApoA-I. In some embodiments, the complex includes ApoA-I having at least 90% sequence identity to SEQ ID NO: 1:

[0128] PPQSPWDRVKDLATVYVDVLKDSGRDYVSQFEGSALGKQLNLKLLDNWDSVTSTFSKLREQLGPVTQEFWDNLEKETEGLRQEMSKDLEEVKAKVQPYLDDFQKKWQEEMELYRQKVEPLRA ELQEGARQKLHELQEKLSPLGEEMRDRARAHVDALRTHLAPYSDELRQRLAARLEALKENGGARLAEYHAKATEHLSTLSEKAKPALEDLRQGLLPVLESFKVSFLSALEEYTKKLNTQ(SEQ ID NO:1).

[0129] In other embodiments, the complex comprises an ApoA-I having at least 95% sequence identity to SEQ ID NO: 1. In other embodiments, the complex comprises an ApoA-I having at least 98% sequence identity to SEQ ID NO: 1. In other embodiments, the complex comprises an ApoA-I having at least 99% sequence identity to SEQ ID NO: 1. In other embodiments, the complex comprises an ApoA-I having 100% sequence identity to SEQ ID NO: 1.

[0130] 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.

[0131] Apolipoprotein molecules can include chimeric apolipoproteins, and the chimeric apolipoproteins include apolipoproteins and one or more connected functional parts, such as one or more CRN-001 complexes, one or more targeting moieties, parts with desired biological activity, affinity tags for auxiliary purification, and / or reporter molecules for characterization or positioning studies. The biologically active connecting part can have an activity that can enhance and / or synergistically enhance the biological activity of the compound incorporated into the complex of the present disclosure. For example, the biologically active part can have antimicrobial (e.g., antifungal, antibacterial, antiprotozoal, antibacterial, antifungal or antiviral) activity. In one embodiment, the connecting functional part of the chimeric apolipoprotein is not in contact with the hydrophobic surface of the complex. In another embodiment, the connecting functional part is in contact with the hydrophobic surface of the complex. In some embodiments, the functional part of the chimeric apolipoprotein can be inherent to natural proteins. In some embodiments, the chimeric apolipoprotein includes a ligand or sequence that is recognized by a cell surface receptor or other cell surface part or can interact with a cell surface receptor or other cell surface part.

[0132] In one embodiment, the chimeric apolipoprotein includes a targeting portion that is not inherent to the natural apolipoprotein, such as a Saccharomyces cerevisiae α-mating factor peptide, folic acid, transferrin, or lactoferrin. In another embodiment, the chimeric apolipoprotein includes a portion with a desired biological activity that enhances and / or synergistically enhances the activity of the compound incorporated into the complex of the present disclosure. In one embodiment, the chimeric apolipoprotein may include a functional portion inherent to the apolipoprotein. An example of an inherent functional portion of an apolipoprotein is an inherent targeting portion formed approximately by amino acids 130-150 of human ApoE, which contains a receptor binding region recognized by a member of the low-density lipoprotein receptor family. Other examples of inherent functional portions of apolipoproteins include an ApoB-100 region that interacts with a low-density lipoprotein receptor and an ApoA-I region that interacts with a type B1 scavenger receptor. In other embodiments, functional portions may be added synthetically or recombinantly to produce chimeric apolipoproteins. Another example is an apolipoprotein with a presequence or presequence from another preproapolipoprotein (e.g., a presequence from preproApoA-II replaces the presequence of preproApoA-I). Another example is an apolipoprotein in which some amphipathic sequence segments have been replaced by other amphipathic sequence segments from another apolipoprotein.

[0133] As used herein, "chimeric" refers to two or more molecules that can exist separately and are connected together to form a single molecule with the desired functions of all its constituent molecules. The constituent molecules of the chimeric molecule can be connected by chemical conjugation synthesis, or, when the constituent molecules are all polypeptides or their analogs, the polynucleotides encoding the polypeptides can be recombinantly fused together so that a single continuous polypeptide is expressed. Such chimeric molecules are called fusion proteins. "Fusion proteins" are chimeric molecules in which the constituent molecules are all polypeptides and are connected (fused) to each other so that the chimeric molecules form a continuous single chain. The various components can be directly connected to each other or can be coupled by one or more linkers. One or more segments of the various components can, for example, be inserted into the sequence of apolipoprotein, or, as another example, can be added to the N-terminus or C-terminus of the sequence of apolipoprotein. For example, the fusion protein may contain an antibody light chain, an antibody fragment, a heavy chain antibody, or a single domain antibody.

[0134] In some embodiments, chimeric apolipoprotein is prepared by chemically conjugating apolipoprotein and the functional part to be connected. Methods of chemically conjugating molecules are well known to those skilled in the art. Such methods will vary according to the structure of the part to be connected, but are easily determined by those skilled in the art. Polypeptides generally contain a variety of functional groups, such as carboxylic acid (-COOH), free amino groups (-NH2) or sulfhydryl (-SH) groups, which can be used to react with suitable functional groups on the functional part or on the joint to bind the part to it. The functional part can be connected to a functional group on the N-terminus, C-terminus or internal residue of the apolipoprotein molecule (i.e., a residue in an intermediate position between the N-terminus and the C-terminus). Alternatively, apolipoprotein and / or the part to be labeled can be derivatized to expose or connect additional reactive functional groups.

[0135] In some embodiments, a fusion protein comprising a functional portion of a polypeptide is synthesized using a recombinant expression system. Typically, this involves generating a nucleic acid (e.g., DNA) sequence encoding apolipoprotein and the functional portion so that the two polypeptides will be in frame when expressed, placing the DNA under the control of a promoter, expressing the protein in a host cell, and isolating the expressed protein.

[0136] Nucleic acids encoding chimeric apolipoproteins can be incorporated into recombinant expression vectors in a form suitable for expression in host cells. As used herein, "expression vectors" are nucleic acids that can be transcribed and translated into polypeptides when introduced into appropriate host cells. 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.).

[0137] In some embodiments, apolipoprotein has been modified so that when apolipoprotein is incorporated into the complex 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 the formation of intramolecular or intermolecular disulfide bonds, such as by site-directed mutagenesis. In another embodiment, a chemical cross-linking agent is used to form intermolecular connections between apolipoprotein molecules to enhance the stability of the complex. Intermolecular cross-linking prevents or reduces the dissociation of apolipoprotein molecules from the complex and / or prevents displacement by endogenous apolipoprotein molecules in the individual to which the complex is applied. In other embodiments, apolipoprotein is modified by chemical derivatization of one or more amino acid residues or by site-directed mutagenesis to confer cell surface receptor targeting ability or to be recognized by cell surface receptors.

[0138] By engineering receptor recognition properties into apolipoproteins, the complex can be targeted to specific cell surface receptors. For example, the complex can be targeted to a specific cell type known to carry a specific type of infectious agent, such as by modifying the apolipoprotein so that it can interact with receptors on the surface of the targeted cell type. For example, the complex can be targeted to macrophages by changing the apolipoprotein to confer recognition of the macrophage endocytic class A scavenger receptor (SR-A). Modifying the apolipoprotein by site-directed mutagenesis to replace one or more positively charged amino acids with neutral or negatively charged amino acids can confer SR-A binding ability to the complex. SR-A recognition can also be conferred by preparing chimeric apolipoproteins, which include N- or C-terminal extensions of ligands recognized by SR-A or amino acid sequences with a high concentration of negatively charged residues. The complex containing apolipoproteins can also interact with apolipoprotein receptors, such as but not limited to ABCA1 receptors, ABCG1 receptors, giant proteins, Cubulin and HDL receptors such as SR-B1.

[0139] 5.1.3.2. Apolipoprotein mimics

[0140] Peptides, peptide analogs and agonists that mimic the activity of apolipoprotein (collectively referred to herein as "apolipoprotein peptide mimetics") can also be combined with one or more other lipid binding proteins, or used alone in the complexes described herein. Non-limiting examples of peptides and peptide analogs corresponding to apolipoproteins and agonists that mimic the activity of ApoA-I, ApoA-IM, ApoA-II, ApoA-IV and ApoE suitable for inclusion in the complexes and compositions described herein are disclosed in US Patent Nos. 6,004,925, 6,037,323 and 6,046,166 (granted to Dasseux et al.), US Patent No. 5,840,688 (granted to Tso), US Patent No. 6,743,778 (granted to Kohno), US Publication No. 2004 / 026 6671, 2004 / 0254120, 2003 / 0171277 and 2003 / 0045460 (to Fogelman), US Publication No. 2006 / 0069030 (to Bachovchin), US Publication No. 2003 / 0087819 (to Bielicki), US Publication No. 2009 / 0081293 (to Murase et al.), and PCT Publication No. WO / 2010 / 093918 (to Dasseux et al.), the disclosures of which are incorporated herein by reference in their entirety. These peptides and peptide analogs may comprise L-amino acids or D-amino acids or a mixture of L and D-amino acids. They may also include one or more non-peptide or amide bonds, such as one or more well-known peptide / amide isosteres. Such apolipoprotein peptide mimetics can be synthesized or made using any technique known in the art for peptide synthesis, including, for example, the techniques described in US Patent Nos. 6,004,925; 6,037,323; and 6,046,166.

[0141] In some embodiments, the lipid binding protein molecule comprises an apolipoprotein peptidomimetic molecule and optionally one or more apolipoprotein molecules, such as those described above.

[0142] In some embodiments, the apolipoprotein peptidomimetic molecule comprises an ApoA-I peptidomimetic, an ApoA-II peptidomimetic, an ApoA-IV peptidomimetic, or an ApoE peptidomimetic, or a combination thereof.

[0143] 5.1.4. Amphiphilic molecules

[0144] An amphiphilic molecule is a molecule that has both a hydrophobic (non-polar) and a hydrophilic (polar) component. Amphiphilic molecules that can be used in the complexes described herein include lipids (e.g., as described in Section 5.1.4.1), detergents (e.g., as described in Section 5.1.4.2), fatty acids (e.g., as described in Section 5.1.4.3), and non-polar molecules and sterols covalently linked to polar molecules, such as, but not limited to, sugars or nucleic acids (e.g., as described in Section 5.1.4.4).

[0145] The complex can include a single type of amphiphilic molecule (e.g., a single type of phospholipid or a mixture of phospholipids) or can contain a combination of multiple types of amphiphilic molecules (e.g., a phospholipid and a detergent). The complex can contain an amphiphilic molecule or a combination of amphiphilic molecules configured to promote the solubilization of the lipid binding protein molecule.

[0146] In some embodiments, the amphiphilic molecules included include a phospholipid, a detergent, a fatty acid, a non-polar moiety, or a sterol, or a combination thereof (eg, selected from the types of amphiphilic molecules discussed above) covalently linked to a sugar.

[0147] In some embodiments, the amphiphilic molecule comprises or consists of a phospholipid molecule. In some embodiments, the phospholipid molecule comprises a negatively charged phospholipid, a neutral phospholipid, a positively charged phospholipid, or a combination thereof. In some embodiments, the phospholipid molecule contributes a net charge of 1-3 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 molecule consists of a combination of negatively charged and neutral phospholipids. In some embodiments, the molar ratio of negatively charged phospholipids to neutral phospholipids is in the range of 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.

[0148] In some embodiments, the amphiphilic molecule comprises a neutral phospholipid and a negatively charged phospholipid in a weight ratio of 95:5 to 99:1.

[0149] 5.1.4.1. Lipids

[0150] The complex based on lipid-binding protein can include one or more lipids. In various embodiments, one or more lipids can be saturated and / or unsaturated, natural and / or synthetic, charged or uncharged, zwitterionic or uncharged. In some embodiments, lipid molecules (e.g., phospholipid molecules) can contribute together to the net charge of each lipid-binding protein molecule 1-3 (e.g., 1-3, 1-2, 2-3, 1, 2 or 3) in the complex. In some embodiments, the net charge is negative. In other embodiments, the net charge is positive.

[0151] In some embodiments, the lipid comprises a phospholipid. The phospholipid can have two identical or different acyl chains (e.g., chains with different carbon atoms, different saturations between acyl chains, different branches of acyl chains, or a combination thereof). The lipid can also be modified to comprise a fluorescent probe (e.g., as described at avantilipids.com / product-category / products / fluorescent-lipids / ). Preferably, the lipid comprises at least one phospholipid.

[0152] The phospholipids can have unsaturated or saturated acyl chains ranging 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 phospholipids used in the complexes of the present disclosure have 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).

[0153] Non-limiting examples of acyl chains present in common fatty acids that may be included in phospholipids are provided in Table 1 below:

[0154]

[0155] The lipids that may be present in the complexes of the present disclosure include, but are not limited to, small alkyl chain phospholipids, egg phosphatidylcholine, soybean phosphatidylcholine, dipalmitoylphosphatidylcholine, dimyristoylphosphatidylcholine, distearoylphosphatidylcholine, 1-myristoyl-2-palmitoylphosphatidylcholine, 1-palmitoyl-2-myristoylphosphatidylcholine, 1-palmitoyl-2-stearoylphosphatidylcholine, 1-stearoyl-2-palmitoylphosphatidylcholine, dioleoylphosphatidylcholine, dioleoylphosphatidylethanolamine, dilauroylphosphatidylglycerolphosphatidylcholine, phosphatidylserine, phosphatidylethanolamine, phosphatidylinositol, phosphatidylglycerol, diphosphatidylglycerol, such as dimyristoylphosphatidylglycerol, dipalmitoylphosphatidylglycerol, distearoylphosphatidylcholine, 1 ...palmitoylphosphatidylcholine, 1- Fatty acylphosphatidylglycerol, dioleoylphosphatidylglycerol, dimyristoylphosphatidic acid, dipalmitoylphosphatidic acid, dimyristoylphosphatidylethanolamine, dipalmitoylphosphatidylethanolamine, dimyristoylphosphatidylserine, dipalmitoylphosphatidylserine, brain phosphatidylserine, brain sphingomyelin, dipalmitoyl sphingomyelin, dipalmitoyl sphingomyelin, egg sphingomyelin, milk sphingomyelin, plant sphingomyelin, distearoyl sphingomyelin, dipalmitoylphosphatidylglycerol salt, phosphatidic acid, galactocerebroside, ganglioside, cerebroside, dilaurylphosphatidylcholine, (1,3)-D-mannosyl-(1,3) diacylglycerol, aminophenyl glycoside, 3-cholesterol-6'-(glycosylthio)hexyl ether glycolipid, and cholesterol and its derivatives. Synthetic lipids, such as synthetic palmitoyl sphingomyelin or N-palmitoyl-4-hydroxysphingosine-1-phosphocholine (a form of plant sphingomyelin) can be used to minimize lipid oxidation.

[0156] In some embodiments, the lipid-binding protein-based complex comprises two types of phospholipids: neutral lipids, such as phosphatidylcholine and / or sphingomyelin (abbreviated as SM), and charged phospholipids (e.g., negatively charged phospholipids). "Neutral" phospholipids have a net charge of about zero at physiological pH. In many embodiments, neutral phospholipids are zwitterions, although other types of net neutral phospholipids are known and can be used. In some embodiments, the molar ratio of charged phospholipids (e.g., negatively charged phospholipids) to neutral phospholipids ranges from 1:1 to 1:3, such as about 1:1, about 1:2, or about 1:3.

[0157] Neutral phospholipids can comprise one or both of for example lecithin and / or SM, and can optionally comprise other neutral phospholipids.In some embodiments, neutral phospholipids comprises lecithin, but does not comprise SM.In other embodiments, neutral phospholipids comprises SM, but does not comprise lecithin.In other embodiments, neutral phospholipids comprises both lecithin and SM.Except lecithin and / or SM, all these specific exemplary embodiments can comprise neutral phospholipids, but do not comprise such extra neutral phospholipids in many embodiments.

[0158] As used herein, the expression "SM" includes sphingomyelin derived from or obtained from natural sources, and analogs and derivatives of naturally occurring SM that are not affected by LCAT hydrolysis, such as naturally occurring SM.SM is a phospholipid that is very similar to lecithin in structure, but unlike lecithin, it does not have a glycerol backbone, and therefore does not have an ester bond connecting an acyl chain. On the contrary, SM has a ceramide backbone, in which an amide bond connects an acyl chain.SM can be obtained, for example, from milk, eggs or brain.SM analogs or derivatives can also be used. The non-limiting examples of useful SM analogs and derivatives include but are not limited to palmitoyl sphingomyelin, N-palmitoyl-4-hydroxysphingosine-1-phosphocholine (a form of plant sphingomyelin), palmitoyl sphingomyelin, stearoyl sphingomyelin, D-erythro-N-16:0-sphingomyelin and its dihydro isomers, D-erythro-N-16:0-dihydrosphingomyelin. Synthetic SMs, such as synthetic palmitoyl sphingomyelin or N-palmitoyl-4-hydroxysphingosine-1-phosphocholine (phytosphingomyelin), can be used to produce a more homogeneous complex with fewer contaminants and / or oxidation products than sphingolipids of animal origin. Methods for synthesizing SMs are described in US Publication No. 2016 / 0075634.

[0159] Sphingomyelin isolated from natural sources can be artificially enriched in a particular saturated or unsaturated acyl chain. For example, milk sphingomyelin (Avanti Phospholipid, Alabaster, Ala) is characterized by long saturated acyl chains (i.e., acyl chains with 20 or more carbon atoms). In contrast, egg sphingomyelin is characterized by short saturated acyl chains (i.e., acyl chains with less than 20 carbon atoms). For example, only about 20% of milk sphingomyelin contains C16:0 (16 carbons, saturated) acyl chains, while 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, or vice versa.

[0160] SM can be semi-synthetic so that it has a specific acyl chain. For example, milk sphingomyelin can be first purified from milk, and then a specific acyl chain, such as a C16:0 acyl chain, can be cut and replaced by another acyl chain. SM can also be completely synthesized, for example, by large-scale synthesis. See, for example, Dong et al., US Patent No. 5,220,043, entitled Synthesis of D-erythro-sphingomyelins, granted on June 15, 1993; Weis, 1999, Chem. Phys. Lipids 102 (1-2): 3-12. SM can be completely synthesized, for example, as described in US Publication No. 2014 / 0275590.

[0161] The length and saturation level of the acyl chain comprising semisynthetic or synthetic SM can be selectively changed.Acyl chain can be saturated or unsaturated, and can contain about 6 to about 24 carbon atoms.Each chain can contain the same number of carbon atoms, or, each chain can contain different numbers of carbon atoms.In some embodiments, semisynthetic or synthetic SM comprises mixed acyl chain, so that one chain is saturated and one chain is unsaturated.In such mixed acyl chain SM, chain length can be the same or different.In other embodiments, the acyl chain of semisynthetic or synthetic SM is all saturated or all unsaturated.Equally, chain can contain the same or different numbers of carbon atoms.In some embodiments, two acyl chains comprising semisynthetic or synthetic SM are identical.In a specific embodiment, chain corresponds to the acyl chain of naturally occurring fatty acid, for example oleic acid, palmitic acid or stearic acid.In another embodiment, use the SM with saturated or unsaturated functionalized chain.In another specific embodiment, acyl chain is all saturated and contains 6 to 24 carbon atoms. Non-limiting examples of acyl chains present in common fatty acids that may be included in semi-synthetic and synthetic SMs are provided in Table 1 above.

[0162] In some embodiments, the SM is a palmitoyl SM, such as a synthetic palmitoyl SM, which has a C16:0 acyl chain, or an egg SM, which includes a palmitoyl SM as a major component.

[0163] In a specific embodiment, a functionalized SM is used, such as a phytosphingomyelin.

[0164] Lecithin can be derived from natural sources or separated, or can be synthesized. The example of suitable lecithin separated from natural sources includes but is not limited to egg phosphatidylcholine and soybean phosphatidylcholine. Other non-limiting examples of suitable lecithin include dipalmitoylphosphatidylcholine, dimyristoylphosphatidylcholine, distearoylphosphatidylcholine, 1-myristoyl-2-palmitoylphosphatidylcholine, 1-palmitoyl-2-myristoylphosphatidylcholine, 1-palmitoyl-2-stearoylphosphatidylcholine, 1-stearoyl-2-palmitoylphosphatidylcholine, 1-palmitoyl-2-oleoylphosphatidylcholine, 1-oleoyl-2-palmitoylphosphatidylcholine, dioleoylphosphatidylcholine and ether derivatives or analogs thereof.

[0165] The lecithin derived from or separated from natural origin can be enriched to include specific acyl chains. In the embodiment adopting semi-synthetic or synthetic lecithin, the characteristic of acyl chain can be selectively changed, as discussed above in conjunction with SM. In some embodiments of complex described herein, two acyl chains on lecithin are identical. In some embodiments of the complex comprising SM and lecithin, the acyl chains of SM and lecithin are all identical. In a specific embodiment, the acyl chain corresponds to myristic acid, palmitic acid, oleic acid or stearic acyl chains.

[0166] The complexes disclosed herein may include one or more negatively charged phospholipids (e.g., alone or in combination with one or more neutral phospholipids). As used herein, "negatively charged phospholipids" are phospholipids with a net negative charge at physiological pH. Negatively charged phospholipids may include a single type of negatively charged phospholipid, or a mixture of two or more different negatively charged phospholipids. In some embodiments, the 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 or potassium salts). In some embodiments, the negatively charged phospholipids include one or more of phosphatidylinositol, phosphatidylserine, phosphatidylglycerol and / or phosphatidic acid. In a specific embodiment, the negatively charged phospholipids include or consist of a salt of phosphatidylglycerol or a salt of phosphatidylinositol. In another specific embodiment, the negatively charged phospholipid comprises or consists of 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] or DPPG or a salt thereof.

[0167] Negatively charged phospholipids can be obtained from natural sources or prepared by chemical synthesis. In embodiments using synthetic negatively charged phospholipids, the characteristics of the acyl chains can be selectively changed, as discussed above in conjunction with SM. In some embodiments of the complexes disclosed herein, the two acyl chains on the negatively charged phospholipids are the same. In some embodiments, the acyl chains of all types of phospholipids contained in the complexes disclosed herein are the same. In a specific embodiment, the complex comprises negatively charged phospholipids and / or SMs all having C16:0 or C16:1 acyl chains. In a specific embodiment, the fatty acid portion of the SM is primarily C16:1 palmitoyl. In a specific embodiment, the acyl chains of the charged phospholipids, phosphatidylcholine and / or SM correspond to the acyl chains of palmitic acid. In another specific embodiment, the acyl chains of the charged phospholipids, phosphatidylcholine and / or SM correspond to the acyl chains of oleic acid.

[0168] Examples of positively charged phospholipids that can be included in the complexes of the present disclosure include N1-[2-((1S)-1-[(3-aminopropyl)amino]-4-[bis(3-amino-propyl)amino]butylcarboxamido)ethyl]-3,4-bis[oleoyloxy]-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- Glycerol-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(oleoyloxy)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)-1propylammonium methylsulfate and salts thereof (e.g., chloride salts or bromide salts).

[0169] The lipids used are preferably at least 95% pure, and / or have reduced levels of oxidants (such as, but not limited to, peroxides). Lipids obtained from natural sources preferably have fewer polyunsaturated fatty acid moieties and / or less oxidizable fatty acid moieties. The oxidation level in a sample can be determined using iodine titration, which provides a peroxide value, expressed as milliequivalents of separated iodine per kilogram of sample, abbreviated as meq O / kg. See, e.g., Gray, 1978, Measurement of Lipid Oxidation: A Review, Journal of the American Oil Chemists Society 55: 539-545; Heaton, FW 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, e.g., 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.

[0170] In some embodiments, the complex may include a small amount of additional lipids. In fact, any type of lipid can be used, including but not limited to lysophospholipids, galactosylceraside, ganglioside, cerebroside, glyceride, triglyceride and sterol and sterol derivatives (e.g., phytosterol, animal sterol such as cholesterol or sterol derivatives such as cholesterol derivatives). For example, the complex of the present disclosure may contain cholesterol or cholesterol derivatives, such as cholesterol esters. Cholesterol derivatives may also be substituted cholesterol or substituted cholesterol esters. The complex of the present disclosure may also contain oxidized sterols, such as but not limited to oxidized cholesterol or oxidized sterol derivatives (e.g., but not limited to oxidized cholesterol esters). In some embodiments, the complex does not include cholesterol and / or its derivatives (e.g., cholesterol esters or oxidized cholesterol esters).

[0171] 5.1.4.2. Detergents

[0172] The complex may contain one or more detergents. The detergent may 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 lauramide hydrochloride, dodecyl ammonium chloride, decyl trimethyl ammonium bromide and hexadecyl trimethyl ammonium sulfate. Exemplary anionic detergents include cholesterol hemisuccinate, cholate, alkyl sulfates and alkyl sulfonates.

[0173] 5.1.4.3. Fatty acids

[0174] The complex may contain one or more fatty acids. The one or more fatty acids may include short chain fatty acids (e.g., butyric acid, isobutyric acid, valeric acid, or isovaleric acid) with an aliphatic tail of 5 or less carbons, medium chain fatty acids (e.g., caproic acid, caprylic acid, capric acid, or lauric acid) with an aliphatic tail of 6 to 12 carbons, long chain fatty acids (e.g., myristic acid, palmitic acid, stearic acid, or arachidic acid) with an aliphatic tail of 13 to 21 carbons, very long chain fatty acids (e.g., behenic acid, lignoceric acid, or cerotic acid) with an aliphatic tail of 22 or more carbons, 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, cedar acid, oleic acid, elaidic acid, vaccenic acid, linoleic acid, trans-linoleic acid, alpha-linolenic acid, arachidonic acid, eicosapentaenoic acid, erucic acid, or docosahexaenoic acid), or a combination 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.

[0175] 5.1.4.4. Sugar-linked nonpolar molecules and sterols

[0176] The complex may contain one or more amphiphilic molecules comprising a non-polar molecule or moiety (e.g., a hydrocarbon chain, an acyl group or a diacyl group) or a sterol (e.g., cholesterol) linked to a sugar (e.g., a monosaccharide such as glucose or galactose, or a disaccharide such as maltose or trehalose). The sugar may be a modified sugar or a substituted sugar. Exemplary amphiphilic molecules comprising a non-polar molecule linked to a sugar include dodec-2-yloxy-β-D-maltoside, tridec-3-yloxy-β-D-maltoside, tridec-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.

[0177] In some embodiments, the non-polar moiety is an acyl or diacyl chain.

[0178] In some embodiments, the sugar is a modified sugar or a substituted sugar.

[0179] 5.1.5. Preparation

[0180] The lipid binding protein-based complex can be formulated according to the intended route of administration, for example according to techniques known in the art (e.g., as described in Allen et al., eds., 2012, Remington: The Science and Practice of Pharmacy, 22nd Edition, Pharmaceutical Press, London, UK).

[0181] CER-001 intended for administration by infusion may be formulated in phosphate buffered saline with sucrose and mannitol excipients, for example as described in WO 2012 / 109162.

[0182] 5.2. Subject population

[0183] The subject that can be treated according to the methods described herein is preferably a mammal, most preferably a human.

[0184] In some aspects, the subject suffers from or is at risk of developing leukocytosis. Leukocytosis can be attributed to various conditions, such as infection, inflammatory processes, and primary bone marrow conditions (e.g., acute leukemia, chronic leukemia, and myeloproliferative conditions). Physical stress, such as seizures, anesthesia, or overexertion, and emotional stress can also increase white blood cell counts. Leukocytosis can also be caused by medications, such as corticosteroids, lithium, and beta agonists. Increased eosinophil or basophil counts caused by various infections, allergic reactions, and other causes can sometimes also lead to leukocytosis.

[0185] In some embodiments, a subject with leukocytosis has a white blood cell count greater than 11 x 10 9 / L and / or less than 100x10 9 / L. In some embodiments, the subject's white blood cell count is 11x10 9 / L and 50x10 9 / L, between 11x10 9 / L and 25x10 9 / L, between 25x10 9 / L and 50x10 9 / L, or between 50x10 9 / L and 100x10 9 / L.

[0186] In some embodiments, the subject has a white blood cell count greater than 11 x 10 9 / L, larger than 12x10 9 / L, greater than 15x10 9 / L or larger than 20x10 9 / L. Alternatively or additionally, in some embodiments, the subject has a white blood cell count of less than 100 x 10 9 / L, less than 50x10 9 / L, less than 25x10 9 / L or less than 15x10 9 / L.

[0187] In some embodiments, the subject has a white blood cell count of 11 x 10 9 / L and 15x10 9 / L, or between 15x10 9 / L and 20x10 9 / L.

[0188] The subject may suffer from one or more symptoms associated with leukocytosis. Exemplary symptoms associated with leukocytosis include fever, bleeding or bruising, sweating, pain or tingling in the legs, arms or abdomen, vision problems (e.g., blurred vision, double vision, blind spots, cloudy vision, dim vision, or a combination thereof), unclear thinking, loss of appetite, or breathing difficulties (e.g., shortness of breath, lower than normal blood oxygen levels).

[0189] In some embodiments, the subject has or is at risk for developing leukocytosis due to inflammation, infection, a leukocyte disorder, physical stress, emotional stress, a drug, or an allergic reaction.

[0190] In some embodiments, the subject having or at risk for developing leukocytosis has an infection, such as a bacterial, fungal, parasitic, or viral infection (e.g., a coronavirus infection such as COVID-19 or influenza).

[0191] In some embodiments, the subject having or at risk for developing leukocytosis has diabetes.

[0192] In some embodiments, the subject having or at risk for developing leukocytosis has received or is currently receiving a medication that has leukocytosis as a side effect, such as a steroid, corticosteroid, lithium, or a beta agonist.

[0193] In some embodiments, the subject having or at risk of developing leukocytosis has a leukocyte disorder.

[0194] In some embodiments, the leukocyte disorder is a primary myeloid disorder (eg, an acute leukemia, a chronic leukemia, or a myeloproliferative disorder).

[0195] In some embodiments, the leukocyte disorder is neutrophilia (eg, idiopathic neutrophilia, which in some embodiments is chronic idiopathic neutrophilia).

[0196] In some embodiments, the leukocyte disorder is hemolytic anemia.

[0197] In some embodiments, the leukocyte disorder is thrombocytopenia (eg, idiopathic thrombocytopenia or essential thrombocytopenia).

[0198] In some embodiments, the leukocyte disorder is cancer (eg, bone cancer or blood cancer).

[0199] In some embodiments, the leukocyte disorder is a myeloproliferative disorder.

[0200] In some embodiments, the leukocyte disorder is a blood cancer.

[0201] In some embodiments, the leukocyte disorder is lymphoma.

[0202] In some embodiments, the leukocyte disorder is a leukemia (eg, lymphocytic leukemia, chronic eosinophilic leukemia, chronic myeloid leukemia, or chronic neutrophilic leukemia).

[0203] In some embodiments, the leukocyte disorder is polycythemia vera.

[0204] In some embodiments, the leukocyte disorder is myelofibrosis, such as primary myelofibrosis.

[0205] In some embodiments of the method of treating a subject having or at risk of developing leukocytosis and / or a subject having one or more symptoms associated with leukocytosis, administration of the lipid binding protein-based complex is effective to reduce the subject's white blood cell count and / or improve one or more symptoms associated with leukocytosis.

[0206] In some aspects, the subject has endothelial dysfunction (e.g., a subject experiencing or having experienced an acute coronary syndrome or stroke). Examples of acute coronary syndromes include myocardial infarction (e.g., ST elevation myocardial infarction, non-ST elevation myocardial infarction), and unstable angina.

[0207] In some embodiments, the subject is experiencing an acute coronary syndrome. In other embodiments, the subject has experienced an acute coronary syndrome. In other embodiments, the subject is experiencing a stroke. In other embodiments, the subject has experienced a stroke.

[0208] In some aspects, the subject suffers from carditis or is at risk of developing carditis (e.g., myocarditis and / or pericarditis). In some embodiments, the subject is male. In some embodiments, the subject is less than 60 years old, less than 50 years old, less than 40 years old, less than 30 years old, less than 20 years old and / or at least 5 years old, at least 8 years old, at least 10 years old, at least 12 years old, at least 15 years old, or at least 18 years old.

[0209] In some embodiments, the subject with carditis or at risk of developing carditis has an infection, such as a viral infection or a bacterial infection. Exemplary viral infections include influenza and coronavirus infections. In some embodiments, the subject has a COVID-19 infection. COVID-19 vaccines, such as mRNA COVID-19 vaccines, are associated with carditis, especially in young men. Therefore, in some embodiments, the subject at risk of developing carditis is a young male (e.g., at least 5, 8, 10, 12, 15 or 18 years old and less than 30 or 20 years old). In some embodiments, the subject has received a vaccine (e.g., a COVID-19 vaccine) before administering a lipid-binding protein-based complex (e.g., 1 to 10 days). In other embodiments, the subject receives a vaccine (e.g., a COVID-19 vaccine) after administering a lipid-binding protein-based complex (e.g., 1 to 10 days). In other embodiments, the subject receives a vaccine (e.g., a COVID-19 vaccine) while administering a lipid-binding protein-based complex (e.g., 1 to 10 days). In other embodiments, the subject receives a vaccine (e.g., a COVID-19 vaccine) while administering a lipid-binding protein-based complex (e.g., on the same day).

[0210] In some embodiments of the disclosed methods, the subject has a SOFA score of 1 to 4 prior to treatment with the lipid binding protein-based complex, such as a score of 1, 2, 3, or 4 (see Vincent et al. 1996, Intensive Care Med, 22: 707-710).

[0211] In some embodiments of the methods of the present disclosure, prior to administration of the lipid binding protein-based complex, the subject has a positive test result as determined by an endotoxin activity assay (EEA) TM ) (Spectral Medical) measured endotoxin activity levels > 0.6 (see, Marshall et al., 2004, J Infect Dis. 190(3):527-34).

[0212] On the other hand, the subject is a subject who needs to reduce the serum level of one or more inflammatory markers, such as a subject with an elevated level 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, one or more cytokines include the aforementioned combination, such as 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) 2, 3, 4, 5, 6, 7 or all 8.

[0213] Dosage regimen

[0214] The methods of the present disclosure generally require multiple administrations of the lipid-binding protein-based complex (e.g., CER-001), such as two to 20 separate doses (e.g., 2 to 16, 2 to 12, 2 to 10, 2 to 8, 2 to 6, 2 to 4, 4 to 20, 4 to 16, 4 to 10, 4 to 6, 6 to 20, 6 to 16, 6 to 10, 6 to 8, 8 to 20, 8 to 16, 8 to 10, 10 to 20, 10 to 16, or 16 to 20 separate doses). In some embodiments, the administration regimen may include four or more doses of the lipid-binding protein-based complex (e.g., CER-001), such as five, six, seven, eight, nine, ten, eleven, twelve, or more than twelve doses.

[0215] In some embodiments, the lipid-binding protein-based complex is administered according to the induction and (optional) consolidation schemes as described in Sections 5.3.1 and 5.3.2, respectively. In some embodiments, the lipid-binding protein-based complex can be administered in a single phase, for example, according to the administration scheme described in this section. In some embodiments, the subject is not treated with a lipid-binding protein-based complex according to a maintenance scheme (e.g., a scheme comprising long-term (e.g., one month or longer) administration of a lipid-binding protein-based complex).

[0216] The administration regimen of the lipid binding protein-based complexes of the present disclosure (eg, CER-001) can last up to one week, one week, or more than one week (eg, two weeks).

[0217] For example, a lipid binding protein-based complex (e.g., CER-001) administration regimen may include administering:

[0218] - Five doses of CER-001 in one week;

[0219] - Six doses of CER-001 in one week;

[0220] - Seven doses of CER-001 in one week;

[0221] - Ten doses of CER-001 over two weeks;

[0222] - Twelve doses of CER-001 over two weeks;

[0223] - Fourteen doses of CER-001 over two weeks.

[0224] In one embodiment, the methods of the disclosure comprise administering seven doses of CER-001 in one week, for example, on days 1, 2, 3, 4, 5, 6, and 7.

[0225] In some embodiments of the methods of the present disclosure, the lipid-binding protein-based complex (e.g., CER-001) is administered daily, for example, at least 5 days, at least 6 days, at least 7 days, or more than 7 days (e.g., daily administration for up to one week or daily administration for up to two weeks). In some embodiments of the methods of the present disclosure, the lipid-binding protein-based complex (e.g., CER-001) is administered multiple times about 12 hours apart (e.g., four to six doses are administered about 12 hours apart). In other embodiments, the lipid-binding protein-based complex (e.g., CER-001) is administered less frequently, for example, every other day, twice a week, three times a week, or once a week.

[0226] In practice, an administration window can be provided, for example, to accommodate slight changes in a dosing regimen of multiple doses per week. For example, a window of ±2 days or ±1 day before and after the dosing date can be used.

[0227] The lipid binding protein-based complex (e.g., CER-001) can be administered in the methods of the present disclosure for a predetermined period of time, such as one week. Alternatively, the lipid binding protein-based complex (e.g., CER-001) can be administered continuously until one or more symptoms of the condition are alleviated or until the serum level of one or more inflammatory markers is reduced, such as to normal levels or relative to the subject's baseline value (e.g., a baseline value measured before starting lipid binding protein-based complex (e.g., CER-001) treatment). Reference or "normal" levels of various inflammatory markers are known in the art. For example, the Mayo Clinic Laboratory Test Catalog (www.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.

[0228] The methods of the present disclosure (e.g., methods for treating a condition or symptom described herein) generally include administering a high dose of a lipid-binding protein-based complex (e.g., CER-001). The high dose can be the sum of multiple individual doses (e.g., two, three, four, five, six, seven, eight, nine, 10, or more than 10 individual doses), for example, administered over multiple days (e.g., three, four, five, six, seven, eight, nine, 10, 11, 12, 13, 14, or 15 days). In some embodiments, individual doses of the high dose are administered daily, twice daily (e.g., about 12 hours apart), or two to three days apart.

[0229] In some embodiments, the high dose is effective to improve one or more symptoms associated with leukocytosis.

[0230] In some embodiments, the high dose is an amount effective to increase the subject's HDL and / or ApoA-I blood levels and / or improve the subject's vascular endothelial function, for example by measuring the levels of circulating vascular cell adhesion molecule 1 (VCAM-1) and / or intercellular adhesion molecule 1 (ICAM-1). In some embodiments, the high dose or individual dose is an amount that increases the subject's HDL and / or ApoA-I levels by at least 25%, at least 30%, or at least 35% 2 to 4 hours after administration.

[0231] In embodiments, the high dose is effective to reduce the subject's circulating VCAM-1 by at least 100 ng / mL, at least 200 ng / mL, at least 300 ng / mL, and / or up to 400 ng / mL within two days of the first administration of the lipid-binding protein-based complex. In embodiments, the high dose is effective to reduce the subject's circulating VCAM-1 by at least 100 ng / mL, at least 200 ng / mL, at least 300 ng / mL, and / or up to 400 ng / mL within five days of the first administration of the lipid-binding protein-based complex.

[0232] In embodiments, the high dose is effective to reduce the subject's circulating ICAM-1 by at least 50 ng / mL, at least 75 ng / mL, at least 100 ng / mL and / or up to 100 ng / mL or up to 125 ng / mL within two days of the first administration of the lipid-binding protein-based complex. In embodiments, the high dose is effective to reduce the subject's circulating ICAM-1 by at least 50 ng / mL, at least 75 ng / mL, at least 100 ng / mL and / or up to 125 ng / mL within five days of the first administration of the lipid-binding protein-based complex.

[0233] In some embodiments, the high dose is an amount effective to reduce the serum level of one or more inflammatory markers (e.g., one or more of IL-6, C-reactive protein, D-dimer, ferritin, IL-8, GM-CSF, MCP1, and TNF-α). In some embodiments, the serum level of one or more inflammatory markers is reduced from an elevated range to a normal range, and / or reduced by at least 20%, at least 40%, or at least 60%.

[0234] In some embodiments, the high dose is effective to reduce the serum level of IL-6 by at least 20pg / mL, at least 30pg / mL, or at least 40pg / mL, and / or up to 40pg / mL within two days of the first administration of the lipid-binding protein-based complex. In some embodiments, the high dose is effective to reduce the serum level of IL-6 by at least 20pg / mL, at least 30pg / mL, or at least 40pg / mL, and / or up to 50pg / mL, or up to 60pg / mL within five days of the first administration of the lipid-binding protein-based complex.

[0235] In some embodiments, the high dose is effective to reduce serum levels of C-reactive protein.

[0236] In some embodiments, the high dose is effective to reduce serum levels of D-dimer.

[0237] In some embodiments, the high dose is effective to reduce the serum level of ferritin. In some embodiments, the high dose is effective to reduce the serum level of ferritin by at least 200 ng / mL, at least 300 ng / mL, or at least 400 ng / mL, and / or up to 700 ng / mL within two days of the first administration of the lipid-binding protein-based complex. In some embodiments, the high dose is effective to reduce the serum level of ferritin by at least 200 ng / mL, at least 300 ng / mL, or at least 400 ng / mL, and / or up to 700 ng / mL within five days of the first administration of the lipid-binding protein-based complex.

[0238] In some embodiments, the high dose is effective to reduce the serum level of interleukin 8 (IL-8). In some embodiments, the high dose is effective to reduce the serum level of IL-8 by at least 100 pg / mL, at least 150 pg / mL, and / or up to 300 pg / mL within two days of the first administration of the lipid-binding protein-based complex. In some embodiments, the high dose is effective to reduce the serum level of IL-8 by at least 100 pg / mL, at least 150 pg / mL, and / or up to 300 pg / mL within five days of the first administration of the lipid-binding protein-based complex.

[0239] In some embodiments, the high dose is effective to reduce serum levels of monocyte chemoattractant protein (MCP) 1 and / or tumor necrosis factor alpha (TNF-α) and / or KIM-1.

[0240] In some embodiments, the high dose is effective to reduce the white blood cell count of the subject. In some embodiments, the high dose is effective to reduce the white blood cell count of the subject by at least 2000 WBC / mL, at least 3000 WBC / mL, at least 4000 WBC / mL, at least 5000 WBC / mL and / or up to 8000 WBC / mL within two days of the first administration of the lipid-binding protein-based complex. In some embodiments, the high dose is effective to reduce the white blood cell count of the subject by at least 2000 WBC / mL, at least 3000 WBC / mL, at least 4000 WBC / mL, at least 5000 WBC / mL and / or up to 6000 WBC / mL within five days of the first administration of the lipid-binding protein-based complex.

[0241] In some embodiments, the high dose is effective to transiently increase serum triglyceride levels, eg, for up to 9 days.

[0242] In some embodiments, the high dose is a dose effective to prevent carditis in a subject at risk for carditis or to reduce the severity of carditis in a subject having carditis or at risk for carditis.

[0243] In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) administered to the subject (e.g., an individual dose that forms a high dose when combined with one or more other individual doses) can be based on protein weight (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) in the range of 4 to 40 mg / kg (e.g., 10 to 20 mg / kg). As used herein, the expression "based on protein weight" means that the dose of the lipid-binding protein-based complex (e.g., CER-001) to be administered to the subject is calculated based on the amount of ApoA-I in the lipid-binding protein-based complex (e.g., CER-001) to be administered and the subject's body weight. For example, a subject weighing 70 kg and who is to receive a 20 mg / kg dose of CER-001 would receive an amount of CER-001 that provides 1400 mg ApoA-I (70 kg x 20 mg / kg).

[0244] In other aspects, the lipid-binding protein-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 (e.g., 600 mg to 4000 mg) per administration (based on protein weight).

[0245] In certain embodiments, the dosage of the lipid binding protein-based complex (eg, CER-001) is 600 mg to 3000 mg, 800 mg to 3000 mg, 1000 mg to 2400 mg, or 1000 mg to 2000 mg (based on protein weight) per administration.

[0246] In some aspects, a high dose of a lipid-binding protein-based complex (e.g., CER-001), such as the sum of multiple individual doses, is 600 mg to 40 g (based on protein weight). In a specific embodiment, the high dose is 3 g to 35 g or 5 g to 30 g (based on protein weight).

[0247] The lipid-binding protein-based complex (e.g., CER-001) is preferably administered as an IV infusion. For example, a stock solution of CER-001 can be diluted in normal saline such as normal saline (0.9% NaCl) to a total volume between 125 ml and 250 ml. In some embodiments, a subject weighing less than 80 kg will have a total volume of 125 ml, while a subject weighing at least 80 kg will have a total volume of 250 ml. In some embodiments, the dose of CER-001 is administered with a total volume of 250 ml. The lipid-binding protein-based complex (e.g., CER-001) can be administered within a time range of one hour to 24 hours. Depending on the needs of the subject, administration can be by slow infusion lasting more than one hour (e.g., up to 2 hours or up to 24 hours), or by rapid infusion of one hour or less, or by a single bolus. In one embodiment, the lipid-binding protein-based complex (e.g., CER-001) is administered within a time period of one hour, for example, using an infusion pump at a fixed rate of 125 ml / hr or 250 ml / hr. In one embodiment, a dose of the lipid binding protein-based complex (eg, CER-001) is administered as an infusion over a 24 hour period.

[0248] 5.3.1. Induction regimen

[0249] In one embodiment, an induction regimen suitable for use in the methods of the present disclosure requires administration of multiple doses of a lipid binding protein-based complex (eg, CER-001) on multiple consecutive days (eg, three consecutive days).

[0250] In some embodiments, the induction regimen suitable for the methods of the present disclosure requires twice daily administration of the lipid binding protein-based complex (e.g., CER-001), for example, twice daily administration on multiple consecutive days. Twice daily administration may include, for example, two doses separated by about 12 hours or a morning dose and an evening dose (which may be separated by more or less than 12 hours).

[0251] In one embodiment, the induction regimen comprises two doses per day of the lipid binding protein-based complex (eg, CER-001) for three consecutive days.

[0252] The therapeutic dose of the lipid-binding protein-based complex (e.g., CER-001) administered by infusion in the induction regimen can be in the range of 4-40 mg / kg (e.g., 4-30 mg / kg) based on 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 foregoing 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 lipid-binding protein-based complex (e.g., CER-001) used in the induction regimen is 5 mg / kg. In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) used in the induction regimen is 10 mg / kg. In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) used in the induction regimen is 15 mg / kg. In some embodiments, the dose of lipid-binding protein-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 lipid-binding protein-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.

[0253] In other aspects, the lipid-binding protein-based complex (e.g., CER-001) can be administered on a unit dose basis. The unit dose used in the induction phase can vary from 300 mg to 4000 mg (e.g., 300 mg to 3000 mg) (based on protein weight) per administration by infusion.

[0254] In certain embodiments, the dose of lipid binding protein-based complex (eg, 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 (based on protein weight) per administration by infusion.

[0255] 5.3.2. Consolidation plan

[0256] Consolidation regimens suitable for use in the methods of the present disclosure entail administration of one or more doses of a lipid binding protein-based complex (eg, CER-001) following an induction regimen.

[0257] In one embodiment, the consolidation regimen comprises administering two doses of a lipid binding protein-based complex (e.g., CER-001). For example, the two doses may be administered about 12 hours apart, or as a morning dose and an evening dose (which may be more or less than 12 hours apart).

[0258] In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) in the consolidation regimen may be administered on day 6 of the dosing regimen starting from the induction regimen on day 1. In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) in the consolidation regimen may be administered on day 4 of the dosing regimen starting from the induction regimen on day 1. In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) in the consolidation regimen may be administered on day 5 of the dosing regimen starting from the induction regimen on day 1. In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) in the consolidation regimen may be administered on day 7 of the dosing regimen starting from the induction regimen on day 1.

[0259] The therapeutic dose of the lipid-binding protein-based complex (e.g., CER-001) administered by infusion in the consolidation regimen can be in the range of 4 mg / kg to 40 mg / kg (e.g., 4 to 30 mg / kg) based on 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 foregoing 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 lipid-binding protein-based complex (e.g., CER-001) used in the consolidation regimen is 5 mg / kg. In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) used in the consolidation regimen is 10 mg / kg. In some embodiments, the dose of the lipid-binding protein-based complex (e.g., CER-001) in the consolidation regimen is 15 mg / kg. In some embodiments, the dosage of lipid-binding protein-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 lipid-binding protein-based complex (e.g., CER-001) at a dosage of 5 mg / kg, 10 mg / kg, 15 mg / kg, or 20 mg / kg in one day.

[0260] In other aspects, the lipid-binding protein-based complex (e.g., CER-001) can be administered on a unit dose basis. The unit dose used in the consolidation phase can vary from 300 mg to 4000 mg (e.g., 300 mg to 3000 mg) (based on protein weight) per administration by infusion.

[0261] In certain embodiments, the dose of the lipid binding protein-based complex (eg, 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 (based on protein weight) per administration by infusion.

[0262] The lipid binding protein-based complex (eg, CER-001) can be administered during the consolidation phase in the same manner as described in Section 5.3, eg, as an IV infusion over one hour.

[0263] 5.4. Combination therapy

[0264] The lipid-binding protein-based complex (e.g., CER-001) can be administered to a subject as a monotherapy or as part of a combination therapy regimen as described herein. For example, a combination therapy can include a lipid-binding protein-based complex (e.g., CER-001) combined with standard care treatment for sepsis or other infections. 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.

[0265] In some embodiments, a subject is treated with a lipid-binding protein-based complex (e.g., CER-001) in combination with fluid replacement therapy. In some embodiments, a subject is treated with a lipid-binding protein-based complex (e.g., CER-001) in combination with an antimicrobial agent. In some embodiments, a subject is treated with a lipid-binding protein-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, a subject is treated with a lipid-binding protein-based complex (e.g., CER-001) in combination with an antiviral agent. In some embodiments, a subject is treated with a lipid-binding protein-based complex (e.g., CER-001) in combination with a blood pressure-raising drug (e.g., norepinephrine or epinephrine).

[0266] In some embodiments, the combination therapy regimen may include one or more anti-IL-6 agents and / or one or more other agents for treating CRS, such as corticosteroids (e.g., methylprednisolone and / or dexamethasone). Exemplary anti-IL6 agents include tocilizumab, siltuximab, olotuzumab, ecitumomab, BMS-945429, sirukumab, levitra monoclonal antibody, and CPSI-2364. In some embodiments, a lipid-binding protein-based complex (e.g., CER-001) is administered in combination with tocilizumab. Subjects who have or have had COVID-19 infection can be treated with a lipid-binding protein-based complex (e.g., CER-001) in combination with one or more additional therapies, such as antibodies from recovered COVID-19 patients, antibodies against the COVID-19 spike protein, one or more antiviral agents (e.g., lopinavir, remdesivir, danoprevir, galisvir, darunavir, ritonavir), chloroquine, hydroxychloroquine, azithromycin, interferon (e.g., interferon alpha or interferon beta, each of which can be pegylated), or a combination thereof.

[0267] In certain embodiments, an antihistamine (eg, diphenhydramine, cetirizine, fexofenadine, or loratadine) may be administered prior to administration of the lipid binding protein-based complex (eg, CER-001). Antihistamines may reduce the likelihood of an allergic reaction. 6. Examples

[0268] 6.1. Example 1: CER-001 therapy in a porcine model of LPS-induced AKI

[0269] The ability of CER-001 to attenuate sepsis-associated AKI was evaluated in a lipopolysaccharide (LPS)-induced AKI porcine model.

[0270] Materials and methods

[0271] The pigs were randomly divided into three groups: LPS (endotoxin-positive pigs, n=3), single-dose CER-001-treated pigs (endotoxin-positive pigs, treated with a single dose of CER-001 20 mg / kg; n=3), and multiple-dose CER-001-treated pigs (endotoxin-positive pigs, treated with two doses of CER-001 20 mg / kg; n=3).

[0272] Sepsis was induced in pigs by intravenous infusion of saline solution containing 300 μg / kg LPS at T0. Pigs treated with single dose CER-001 and pigs treated with multiple doses of CER-001 received a 20 mg / kg dose of CER-001 at T0. Pigs treated with multiple doses of CER-001 received a second 20 mg / kg dose of CER-001 three hours later (T3). Serum IL-6, LPS, MCP-1, sVCAM-1, and sICAM-1 levels were monitored over time. Renal tissue damage and fibrosis were assessed at the end of the study period.

[0273] Results

[0274] Increased survival was observed in both CER-001 treated groups compared with the LPS group (data not shown). LPS injection resulted in a time-dependent increase in IL-6 in endotoxemic animals compared with basal conditions (T0). Figure 1 ). CER-001 treatment was able to reverse the effects of LPS, as shown by decreased IL-6 levels ( Figure 1 The second infusion of CER-001 three hours after the first dose (T3) significantly reduced IL-6 serum levels to basal levels at the end of the study (Tend) ( Figure 1 Similarly, high levels of MCP-1 were observed in endotoxemic pigs relative to basal conditions, whereas MCP-1 levels were lower in pigs treated with CER-001 (data not shown).

[0275] Endothelial dysfunction was assessed by measuring sVCAM-1 and sICAM-1 serum levels. A time-dependent increase in sVCAM-1 and sICAM-1 was observed in endotoxemic animals, whereas CER-001 treatment strongly reduced sVCAM-1 and sICAM-1 levels in both treatment groups (95% and 113%, respectively). Figure 2 and Figure 3 ). Consistent with the IL-6 results, two doses of CER-001 ( Figure 2 , "40MG") was more effective in reducing sVCAM-1 to basal levels. In CER-001-treated animals, LPS levels were significantly reduced ( Figure 4 , "20MG" and "40MG"), and the effect was more pronounced after the second infusion of CER-001 ( Figure 4 , “40MG”).

[0276] Endotoxemic renal biopsies showed tubular vacuolization, epithelial flattening, and some apoptotic tubular cells. CER-001 treatment significantly reduced the inflammatory process and tubular damage. In endotoxemic animals, Masson's trichrome staining showed extensive collagen deposition at the interstitial level. In both CER-001-treated groups, collagen deposition in the renal parenchyma was significantly less compared with the LPS group.

[0277] These preclinical data demonstrate that CER-001 treatment reduces systemic inflammation and endothelial dysfunction, thereby limiting renal injury in a porcine model of LPS-induced AKI.

[0278] 6.2. Example 2: A randomized pilot study comparing short-term CER-001 infusions of different doses for preventing sepsis-induced acute kidney injury

[0279] Currently, there are no approved treatments for sepsis-associated AKI. Given that the inflammatory response to endotoxemia is the primary cause of hemodynamic instability and progression to AKI in patients with sepsis, the primary objective of this study is to investigate whether the use of different doses of CER-001 in combination with standard of care (SOC) therapy is safe and effective, providing a new strategy for treating patients with sepsis, reducing the inflammatory response, and preventing progression to AKI. Without being bound by theory, the expected mechanism of action is twofold, including the binding of CER-001 to endotoxin and the direct anti-inflammatory effects of CER-001.

[0280] 6.2.1. Research plan

[0281] Study population: This was a single-center, randomized, dose-ranging (Phase II) study that included patients admitted to the intensive care unit (ICU) of participating centers with sepsis due to intra-abdominal infection or urosepsis. The investigators ensured that all patients who met the following inclusion and exclusion criteria were included in the study.

[0282] Inclusion criteria:

[0283] - Male or non-pregnant female adults ≥18 years old at the time of enrollment;

[0284] - Meet sepsis 3 criteria, defined as an acute increase of at least 2 points in the SOFA score relative to the admission SOFA score;

[0285] - Endotoxin levels (measured by endotoxin activity assay (EEA) TM )Measurement; Spectral

[0286] Medical)>0.6 (see, Marshall et al., 2004, J Infect Dis. 190(3):527-34);

[0287] - Informed consent signed and dated by the patient or legal representative.

[0288] Exclusion criteria:

[0289] - Patients weighing more than 100 kg;

[0290] - Alanine aminotransferase / aspartate aminotransferase (ALT / AST) > 5 times the upper limit of normal;

[0291] - Stage 4 severe chronic kidney disease or requiring dialysis (i.e. estimated glomerular filtration rate (eGFR) <30 ml / min / 1.73 m 2 );

[0292] - WBC <2.0×10^9;

[0293] - Pregnancy or breastfeeding;

[0294] - Received an organ transplant in the past year;

[0295] - Expected to be transferred to another non-study center hospital within 72 hours;

[0296] - Terminal illness, including metastatic disease or hematologic malignancy, with a life expectancy of less than 30 days (assessed by the attending physician) or has been classified as "unrecoverable";

[0297] - Past medical history of end-stage chronic organ failure;

[0298] - Diagnosed with HIV;

[0299] - Uncontrolled bleeding in the last 24 hours;

[0300] - Patients who have used study drug or device within 30 days of the first dose of CER-001.

[0301] Number of subjects: Twenty subjects were recruited and randomly divided into four experimental groups (1:1:1:1): Group A patients continued to receive conventional therapy; Group B: patients added CER-001 5 mg / kg BID on the basis of conventional therapy for 3 days, followed by 5 mg / kg BID on the 6th day; Group C: patients added CER-001 10 mg / kg BID on the basis of conventional therapy for 3 days, followed by 10 mg / kg BID on the 6th day; Group D patients added CER-001 20 mg / kg BID on the basis of conventional therapy for 3 days, followed by 20 mg / kg BID on the 6th day ( Figure 5 ).

[0302] Study duration:This study was completed in 24 weeks (6 months). The enrollment period was approximately 20 weeks (5 months) from the enrollment of the first subject. The end of the study was the last visit of the last subject.

[0303] Primary endpoint: The primary endpoint of the study is to determine the safety and optimal dose of CER-001 in combination with standard of care in patients with persistent sepsis due to Gram-negative bacteria.

[0304] Secondary endpoints: Secondary endpoints were:

[0305] - Changes in endotoxin and IL-6 levels from baseline to days 3, 6, and 9.

[0306] -Baseline was defined as the last measurement performed before dosing on Day 1.

[0307] - Change in SOFA score from baseline to day 3, day 6 and day 9 (Vincent et al. 1996, Intensive Care Med, 22:707–710).

[0308] - Changes from baseline to Day 3, Day 6, and Day 9 in key inflammatory markers (CRP, D-dimer, ferritin, IL-8, GM-CSF, MCP1, and TNF-α).

[0309] -According to the KDIGO criteria (Kidney Disease Improving Global Outcomes.

[0310] KDIGO Clinical Practice Guideline for Acute Kidney Injury. Kidney International Supplements 2012;2:1–138), changes in AKI biomarkers and the onset of AKI.

[0311] - Mortality at day 30.

[0312] - Independent medical experts review outcome data during the trial.

[0313] Intervention / Exposure:Twenty patients who met the eligibility criteria signed and dated the informed consent form approved by the ethics committee (EC) and were randomized and allocated in a ratio (1:1:1:1) to conventional therapy (group A), low-dose CER-001 (group B), medium-dose CER-001 (group C), or high-dose CER-001 (group D). Conventional therapy was adjusted according to clinical conditions. During the patient's participation in this study, concomitant administration of all non-experimental treatments was allowed: any medication taken by the patient (except the study drug specified in the protocol) was considered a concomitant medication and was recorded in the study record.

[0314] Each patient was identified by a patient number at screening. Once assigned to a patient, the patient number was not reused. The randomization list was generated and the assigned sequence was assigned without the involvement of the enrolling investigator. The randomization list was fully concealed to prevent attempts to subvert the randomization.

[0315] Treatment Group: All patients received conventional therapy. The treatment group received additional treatment with the study drug. Specifically:

[0316] - Group A: conventional therapy (i.e., antibiotic treatment and hemodynamic support according to the patient's condition).

[0317] - Group B: conventional therapy + CER-001 5 mg / kg BID for 3 consecutive days, followed by 5 mg / kg BID on day 6.

[0318] - Group C: conventional therapy + CER-001 10 mg / kg BID for 3 consecutive days, followed by 10 mg / kg BID on day 6.

[0319] - Group D: conventional therapy + CER-001 20 mg / kg BID for 3 consecutive days, followed by 20 mg / kg BID on day 6.

[0320] Prior to each CER-001 dose, patients were pretreated with an antihistamine (e.g., dexchlorpheniramine 5 mg or hydroxyzine 100 mg) to avoid any potential infusion reactions. Patients may interrupt or discontinue study drug treatment if any of the following occur:

[0321] Any drug-related adverse event or other reason that the investigator believes jeopardizes the patient's participation in the trial or the interpretation of trial data (e.g., serious intercurrent illness requiring additional nursing measures or preventing further dosing); serious tolerability problems.

[0322] When study medication was discontinued, the study center recorded the reason for discontinuation. Patients were followed up clinically and attempts were made to restart study medication within 2 days of discontinuation if there were no other contraindications.

[0323] Reasons for withdrawal from study medication may include, but are not limited to, the following:

[0324] - Investigator's request, for safety reasons, such as serious adverse reactions;

[0325] - Investigator's request, for other reasons, such as patient non-compliance;

[0326] - Patient's request, for tolerability reasons;

[0327] - Patient's request, for other reasons, such as withdrawal of informed consent.

[0328] Discontinuation of study drug alone does not constitute discontinuation or withdrawal from the study. Patients will continue to be followed as if they had completed the treatment period. Patients who discontinue study drug prematurely (e.g., before completing the third dose) should receive an end-of-study evaluation if possible.

[0329] Statistical analysis: Comparisons between groups were performed using appropriate statistical tests: dichotomous variables (baseline characteristics, mortality, development of AKI) were compared by using chi-square or Fisher's exact test, and continuous baseline characteristics were compared by ANOVA or Kruskall-Wallis test, tStudent or Mann-Whitney U test, as appropriate. Changes in inflammatory markers between groups were compared by ANOVA and presented graphically. The proportion of patients with AKI and mortality were calculated for each group. All analyses were performed using SPSS12.0 for Windows; p < 0.05 was considered statistically significant.

[0330] program: The following procedures were performed during the Screening Visit. After randomization, subjects started treatment within 2 working days.

[0331] -Informed consent

[0332] - Medical history - includes: recording of past and present illnesses, collection of subject demographic data (date of birth, sex, and race).

[0333] -Physical examination, including review of systems, height and weight, BMI, and waist circumference

[0334] - Vital signs (pulse, blood pressure, and oral, aural, axillary, or core temperature).

[0335] - Review of inclusion / exclusion criteria.

[0336] - Adverse events were recorded starting from the time of obtaining informed consent.

[0337] - Collect past medications for the 4 weeks prior to the first dose of the test article. Record all current medications.

[0338] - Complete blood count (CBC) - includes white blood cell count (WBC) with differential, platelet count, red blood cell count (RBC), hemoglobin (Hb), hematocrit (Hct).

[0339] -Fasting chemistry panel / electrolytes: including sodium, potassium, chloride, blood urea nitrogen (BUN; or urea), serum creatinine, calculated creatinine clearance (CKD-EPI), glucose, calcium, phosphorus, total protein, uric acid, AST, ALT, gamma-GT, ALP, total and direct bilirubin, albumin, total cholesterol, HDL, LDL, triglycerides, LDH, CPK,

[0340] -ABG (to evaluate for respiratory and / or metabolic disturbances)

[0341] -ApoA-I (for pharmacokinetic and pharmacodynamic assessments)

[0342] -Coagulation tests - including prothrombin time (PT) (expressed as international normalized ratio [INR]) and partial thromboplastin time (PTT).

[0343] -Urinalysis - includes assessment of specific gravity, pH, protein / albumin, glucose, ketones, and hemoglobin / blood.

[0344] - Microalbuminuria and proteinuria g / 24h

[0345] - Serum or urine pregnancy test within 7 days prior to randomization (for females of childbearing potential).

[0346] -Pharmacokinetic and pharmacodynamic assessments including apoA-I and total cholesterol levels.

[0347] - Use EAA TM Kit to measure endotoxin levels. The kit measures AKI biomarkers (TIMP-2 and IGFBP-7). Inflammatory markers include: CRP, D-dimer, ferritin, IL-6, IL-8, GM-CSF, MCP1, and TNF-α.

[0348] In addition to biological samples collected for routine laboratory evaluations performed on a daily basis at the central laboratory, biological samples are collected for research purposes and include:

[0349] -2 tubes of 5ml serum

[0350] -1 tube of 3ml plasma

[0351] - Urine 30ml

[0352] These samples were used to evaluate additional inflammatory cytokines and urinary biomolecules to obtain a more comprehensive profile of the enrolled patients, better assess the response to treatment, provide more information in follow-up, and more importantly, discover new potential biomarkers that can be used for the early diagnosis of sepsis-induced AKI by ELISA tests and protein array analysis.

[0353] During treatment visits (treatment period): The treatment period was defined as the start of treatment. Visits were scheduled on Days 3, 6, and 9. The final visit was scheduled on Day 30. The following procedures were performed during the treatment visits:

[0354] - Record adverse events and concomitant medications

[0355] - Review appropriate laboratory information

[0356] -Physical examination

[0357] -Assess vital signs (pulse, blood pressure, and oral, aural, axillary, or core temperature)

[0358] - Continuous recording of adverse events and concomitant medications

[0359] - Complete blood count (CBC) - includes white blood cell count (WBC) with differential, platelet count, red blood cell count (RBC), hemoglobin (Hb), hematocrit (Hct).

[0360] -Fasting chemistry panel / electrolytes: including sodium, potassium, chloride, blood urea nitrogen (BUN; or urea), serum creatinine, calculated creatinine clearance (CKD-EPI),

[0361] - Glucose, calcium, phosphorus, total protein, uric acid, AST, ALT, γGT, ALP, total and direct bilirubin, albumin, total cholesterol, HDL, LDL, triglycerides, LDH, CPK

[0362] -ABG (to evaluate for respiratory and / or metabolic disturbances)

[0363] -ApoA-I (for pharmacokinetic and pharmacodynamic assessments)

[0364] -Coagulation tests - including prothrombin time (PT) (expressed as international normalized ratio [INR]) and partial thromboplastin time (PTT).

[0365] -Urinalysis - includes assessment of specific gravity, pH, protein / albumin, glucose, ketones, and hemoglobin / blood.

[0366] - Microalbuminuria and proteinuria g / 24h

[0367] - Serum or urine pregnancy test within 7 days prior to randomization (for females of childbearing potential).

[0368] -Pharmacokinetic and pharmacodynamic assessments will include apoA-I and total cholesterol levels.

[0369] - Use EAA TM Kit to measure endotoxin levels. The kit measures AKI biomarkers (TIMP-2 and IGFBP-7). Inflammatory markers include: CRP, D-dimer, ferritin, IL-6, IL-8, GM-CSF, MCP1, and TNF-α.

[0370] In addition to biological samples collected for routine laboratory evaluations performed in the central laboratory, biological samples were collected for research purposes and included

[0371] -2 tubes of 5ml serum

[0372] -1 tube of 3ml plasma

[0373] - Urine 30ml

[0374] Clinical scores included the SOFA score (Table 2) and the KDIGO criteria for AKI assessment and staging (Table 3). The individual components of each score were recorded.

[0375] Table 2. Sequential Organ Failure Assessment (SOFA) score

[0376]

[0377] MAP = mean arterial pressure; CNS = central nervous system; SaO 2 = peripheral arterial oxygen saturation a Vasoactive drugs (dopamine and norepinephrine μg / kg / min) administered for at least 1 hour

[0378]

[0379] Table 4 provides an overview of the study protocol for this example.

[0380]

[0381]

[0382] Safety evaluation: Safety evaluation was obtained using information collected from the following assessments: physical examination (including weight), vital signs (blood pressure, pulse, temperature), CBC differential, platelet count, blood chemistry and fasting lipid profile [including HDL-cholesterol, LDL-cholesterol and lipoprotein (a)], urea, glucose, 24-hour urine protein measurement, serum creatinine and calculated creatinine clearance (CKD-EPI), and adverse event monitoring. All women of childbearing potential underwent a qualitative serum pregnancy test during the pre-study screening / baseline assessment and subsequently (if clinically indicated). Patients were monitored for the occurrence of adverse events throughout the study and adverse events were recorded. Adverse events voluntarily reported by the subject or discovered as a result of general questioning or physical examination by the investigator were recorded. The duration (start and end date), severity, cause and relationship to the study drug, patient outcome, measures taken, and assessment of the severity of the event were recorded for each reported adverse event.

[0383] Adverse events: definition

[0384] The term "adverse event" is synonymous with the term "adverse experience" used by the FDA. An adverse event (AE) is any unfavorable, undesirable, unplanned clinical occurrence in the form of a sign, symptom, illness, or laboratory or physiological observation, regardless of causality, that occurs in a person participating in a clinical investigation. This includes the following:

[0385] - Any clinically significant worsening of pre-existing condition.

[0386] - Any recurrence of pre-existing condition.

[0387] - AEs resulting from an overdose (whether accidental or intentional) of the investigator's test article (i.e., a dose higher than that prescribed by a healthcare professional for clinical reasons).

[0388] - AEs resulting from misuse of the investigator-supplied test article (i.e., use without clinical justification).

[0389] - AEs related to discontinuation of investigator-supplied test article.

[0390] Surgery is not an AE, but the cause of surgery may be an AE.

[0391] A "pre-existing condition" is a clinical condition (including conditions being treated) that was diagnosed and recorded as part of the subject's medical history before the subject signed the informed consent form. Questions about whether the condition was present before the start of the active phase of the study and whether it increased in severity and / or frequency were used to determine whether an event was a treatment-emergent adverse event (TEAE). An AE was considered treatment-emergent if (1) the AE was a chronic condition that was not present and not part of the subject's medical history at the start of the active phase of the study, or (2) it was present or part of the subject's medical history at the start of the active phase of the study but increased in severity or frequency during the active phase. The active phase of the study began at the first dose of the drug.

[0392] A “serious adverse event” is any AE occurring at any dose that meets one or more of the following criteria:

[0393] - Causes death

[0394] - Life-threatening (see below)

[0395] - Requirement for subject to be hospitalized or prolongation of existing hospitalization (see below)

[0396] - Causes persistent or substantial disability or incapacity (see below)

[0397] - Lead to new malignancies

[0398] - Cause congenital anomalies or birth defects

[0399] Additionally, important medical events that may not result in death, be life-threatening, or require hospitalization may be considered SAEs when, based on appropriate medical judgment, they could endanger the subject and may require medical or surgical intervention to prevent one of the above outcomes. Examples of such events include allergic bronchospasm requiring intensive treatment in the emergency room or at home, blood dyscrasias or seizures that do not require hospitalization, or the development of drug dependence or drug abuse.

[0400] A “life-threatening adverse event” is any AE that, at the time of the event, places the subject at an immediate risk of death. Life-threatening events do not include events that could result in death if they occurred in a more severe form, but that do not pose an immediate risk of death when they actually occur. For example, drug-induced hepatitis that resolves without evidence of liver failure would not be considered life-threatening, although drug-induced hepatitis of a more severe nature could be fatal.

[0401] Hospitalization or prolonged hospital stay is considered a criterion for AE severity. In the absence of an AE, investigators should not report hospitalization or prolonged hospital stay on the form. This would occur if a protocol-required procedure necessitated hospitalization or prolonged hospital stay. Protocol-required daytime or evening investigation visits were not considered severe.

[0402] Reporting time of serious adverse events: Any SAE, regardless of causality, should be reported promptly (no later than 24 hours after the Investigator becomes aware of the SAE) to the Medical Monitor by fax on a completed Serious Adverse Event Form. The Investigator is to report the completed Serious Adverse Event Form to the Medical Monitor (or designee) by fax within 24 hours of receiving follow-up information related to the SAE. Observe and monitor subjects carefully until the condition resolves or stabilizes or its cause is identified. Report any emergency situations promptly (within 24 hours) to the Medical Monitor (or designee) by contacting the Medical Monitor.

[0403] Reportable Events / Information: AEs or SAEs may occur from the time the subject signs the informed consent form to 15 days after the subject's last dose and are not related to the test article or regimen. This includes events that occur during the screening and placebo run-in periods. All AEs and SAEs are recorded in the source documents and documented in the CRF. All AEs and SAEs that occur after the screening period are recorded on the CRF.

[0404] For SEA: The investigator should promptly provide the medical monitor with all documents related to the event (e.g., additional laboratory tests, consultation reports, discharge summaries, autopsy reports, etc.). Submit reports related to the subject's subsequent treatment to the investigator until the event resolves, or in the case of permanent injury, until the condition stabilizes.

[0405] The following events are recorded and reported following the same process as the SAE, within the same timeframe as the SAE:

[0406] Test article abuse and overdose (i.e., use for non-clinical reasons), with or without AEs. An overdose is defined as a dose higher than that prescribed by a healthcare professional for clinical reasons. The decision of whether a dose is an overdose is made by the investigator.

[0407] Unintentional or accidental exposure to the test article, with or without AEs.

[0408] Post-study test article-related SAEs.

[0409] SAEs occurring after unauthorized or accidental use in subjects not participating in the study.

[0410] Abnormal biological or vital sign values ​​that are considered clinically relevant by the Investigator. These events are reported within the same time frame and following the same process as AEs or SAEs.

[0411] Records and Reports: At each required study visit, all AEs that occurred since the last visit were recorded in the Adverse Event Record on the subject's CRF. The information recorded was based on signs or symptoms detected during the subject's physical examination and clinical evaluation. In addition to information obtained from these sources, the subject was asked the following non-specific question: "How have you felt since your last visit?" Signs and symptoms were recorded using standard medical terminology. The Health Outcomes Assessment Survey administered to study subjects was designed to explore the subject's own perception of their quality of life. However, the investigator reviewed the investigation for potential AEs or SAEs and considered the subject's perspective when the AE or SAE was determined to have occurred. The subject's assessment was not influenced by the clinical investigator. Every effort was made to maintain an unbiased assessment. The following AE information was included, if applicable: the specific condition or event and the direction of change; whether the condition was pre-existing (i.e., an acute condition or history of a chronic condition present at the start of the study), and if so, whether it worsened (in severity and / or frequency); date and time of occurrence; severity; causal relationship to the test article; actions taken; and outcome. Any laboratory abnormality considered clinically significant by the investigator was reported as an AE.

[0412] The causal relationship between an AE and the test article was determined by the investigator based on his / her clinical judgment and the following definitions:

[0413] - Definitely Related: The event can be fully explained by the administration of the test article.

[0414] - Probably Related: The event is most likely explained by the administration of the test article rather than by the subject's clinical status or other medications / therapies.

[0415] - Possibly Related: The event may be explained by the administration of the test article or the subject's clinical status or other medications / therapies.

[0416] - Highly Probable Unrelated: The event is most likely explained by the subject's clinical status or other drugs / therapies rather than the test article.

[0417] - Definitely unrelated: The event can be adequately explained by the subject's clinical status or other medications / therapies.

[0418] When evaluating the relationship between test article administration and AEs, the following factors should be considered:

[0419] - The temporal relationship between test article administration and the AE

[0420] - Biological plausibility of the relationship

[0421] - Subject's baseline clinical status or concomitant medications and / or therapies

[0422] Where applicable, whether the AE is alleviated after discontinuation of the test article

[0423] Where applicable, whether the AE that recurs after repeated exposure to the test article is an SAE that is not related to the test article, but which the Investigator or Medical Monitor (or designee) may consider to be related to the conduct of the clinical study, i.e., to the subject's participation in the study. For example, a protocol-related SAE may be an event that occurs during the washout period or is related to a protocol-required procedure. The severity of the AE is assessed according to the National Cancer Institute (NCI) Common Toxicity Criteria for Adverse Events (CTCAE), version 5.0. The following definitions are used for toxicities not defined in the NCI CTCAE:

[0424] - Mild (Grade 1): The AE was noted by the subject but did not interfere with usual activities. This AE did not require discontinuation or dose reduction of the test article.

[0425] -Moderate (Grade 2): AE interferes with usual activities but responds to symptomatic therapy or rest.

[0426] This AE may require a dose reduction but does not require cessation of test article administration.

[0427] - Severe (Grade 3): Despite symptomatic therapy, the AE significantly limits the subject's ability to perform usual activities. In addition, the AE leads to discontinuation or reduction of the test article dose.

[0428] - Life-threatening (Grade 4): AE requires discontinuation of test article administration. The subject is at risk of immediate death.

[0429] Results

[0430] A first cohort of 10 subjects has been treated.

[0431]

[0432] Results from the first cohort are presented in Figures 8A-8J middle.

[0433] Compared with standard of care therapy, CER-001 rapidly improved biomarkers of inflammation, leukocytosis, and endothelial dysfunction, preventing subjects from deteriorating into acute kidney injury. CER-001 treatment was well tolerated at all dose levels (5, 10, and 20 mg / kg, twice daily). No serious treatment-related side effects were observed in this critically ill patient population.

[0434] 6.2.2.1.VCAM and ICAM

[0435] The study is continuing with a total of 20 subjects, n=5 per treatment group. Figure 9A-9F Shown are the changes in VCAM measured by ELISA for the standard care group and the CER-001 group. Statistically significant differences were assessed by using mixed model ANOVA (ns: p>0.05). Fig.9A : Changes from baseline in VCAM in the SOC group and the combined CER-001 group. Fig. 9B : Changes from baseline in VCAM in the SOC group and each study group. Fig. 9C : Change in VCAM for SOC and combined CER-001 groups, reported as percentage relative to peak VCAM levels (peak = 100%). Treatment x study day effect relative to peak p < 0.0001. Fig.9D : Changes in VCAM in the SOC and combined CER-001 groups, according to whether the subjects were enrolled from the ICU or the nephrology department of the center. Fig.9E : Change from baseline in VCAM per subject in the SOC group and the combined CER-001 group. Fig.9F : Change from baseline in VCAM for each subject in each group.

[0436] In general, regimens that delivered CER-001 in addition to SOC reduced VCAM more than SOC alone. The results are summarized in the table below:

[0437] Treatment Effects overall Day 3 Day 6 Day 9 CER-001 and SOC p＝0.0090 NS p＝0.0520 p＝0.0510 Treatment Group p＝0.0870 SOC and 5 NS p＝0.0687 p＝0.0805 SOC and 10 NS p＝0.0556 p＝0.0558 SOC and 20 NS p＝0.0646 NS 5 and 10 NS NS NS 5 and 20 NS NS NS 10 vs 20 NS NS NS

[0438] Figures 10A-10F Shown are the changes in ICAM of the standard care group (SOC group) and the experimental group (CER-001 group) measured by ELISA. Statistically significant differences were assessed by using mixed model ANOVA (ns: p>0.05). Fig. 10A : Changes from baseline in ICAM in the SOC group and the combined CER-001 group. Fig. 10B : Changes from baseline in ICAM in the SOC group and each study group. Fig. 10C : Changes in ICAM for SOC and combined CER-001 groups, expressed as percentage relative to peak ICAM levels (peak = 100%). Treatment x study day effect relative to peak p < 0.0001. Fig. 10D : Changes in ICAM in the SOC and combined CER-001 groups, according to whether the subjects were enrolled from the ICU or the nephrology department of the center. Fig.10E : Change from baseline in ICAM per subject in the SOC group and the combined CER-001 group. Fig.10F: Change from baseline in ICAM for each subject in each study group.

[0439] In general, regimens that delivered CER-001 in addition to SOC reduced ICAM more than SOC alone. The results are summarized in the table below:

[0440] Treatment Effects overall Day 3 Day 6 Day 9 CER-001 and SOC p＝0.0015 NS p＝0.0020 p＝0.0003 Treatment Group p＝0.0127 SOC and 5 p＝0.0552 p＝0.0056 p＝0.0454 SOC and 10 NS p＝0.0213 p＝0.0329 SOC and 20 p＝0.0831 p＝0.0087 p＝0.0023 5 and 10 NS NS NS 5 and 20 NS NS NS 10 vs 20 NS NS NS

[0441] 6.2.2.2. Leukocytes

[0442] Figures 18A-18F The changes in white blood cell counts in the standard of care (SOC) group and the CER-001 group are shown. Fig.18A : Changes from baseline in white blood cell counts in the SOC group and the combined CER-001 group. Fig.18E Shows Fig.18A The individual data points are summarized in . Fig.18B : Changes from baseline in white blood cell counts in the SOC group and each CER-001 group. Fig.18F Shows Fig.18B The individual data points are summarized in . Fig.18C : Change in white blood cell count for SOC and combined CER-001 groups, reported as a percentage relative to peak white blood cell count (peak = 100%). Treatment x study day effect relative to peak p = 0.5492. Fig.18D : Changes in white blood cell counts in the SOC group and the combined CER-001 group, according to whether the subjects were enrolled from the ICU or the nephrology department of the center. Fig.18A , Fig.18B and Fig.18D The unit is cells / μl.

[0443] 6.3. Example 3: CER-001 therapy for the treatment of CRS secondary to Covid-19 infection

[0444] COVID-19 infects host cells through the binding of the viral spike protein (SARS-2-S) to the cell surface receptor angiotensin-converting enzyme 2 (ACE2), and HDL scavenger receptor type B1 (SR-B1) promotes ACE2-dependent viral entry. (Wei et al., Nature Metabolism doi.org / 10.1038 / s42255-020-00324-0). Without being bound by theory, it is believed that lipid-binding protein-based complexes, such as CER-001, can provide therapeutic benefits (e.g., reducing the severity and / or duration of CRS) in subjects with COVID-19 infection by competitively binding to SR-B1, thereby limiting the ability of the virus to infect additional cells.

[0445] A pilot study was conducted to investigate the safety and efficacy of seven CER-001 infusions in patients with CRS secondary to COVID-19 infection. The study consisted of 9 visits:

[0446] Pre-dose (baseline) visit: Assessment of baseline inflammatory markers and safety laboratories.

[0447] • Dosing Visits: 7 doses (Doses 1 to 7) are administered as once daily infusions for 7 days. IL-6 is measured daily from pre-infusion samples.

[0448] Follow-up visit: Patients had a final assessment on Day 8. Inflammatory markers and safety labs were measured.

[0449] The research flow chart is as follows Figure 6 shown.

[0450] 6.3.1. Selection of study subjects

[0451] 6.3.1.1. Inclusion criteria

[0452] Eligible patients who met the following criteria were enrolled in the study:

[0453] 1. Male or non-pregnant female adults aged ≥18 years at the time of enrollment.

[0454] 2. Laboratory-confirmed novel coronavirus (COVID-19) infection identified by polymerase chain reaction (PCR) or other commercial or public health assay in oropharyngeal or anal specimen within 72 hours prior to hospitalization.

[0455] 3. Illness of any duration and at least one of the following:

[0456] a. Radiographic infiltration (chest x-ray, CT scan, etc.), or

[0457] b. Clinical assessment (evidence of rales / crackles on physical examination) and room air

[0458] SpO2 ≤ 93%, or

[0459] c. Requires mechanical ventilation and / or supplemental oxygen, or

[0460] d. Fever that has persisted for the past 24 hours and has not responded to NSAIDs or steroids

[0461] 4. Serum IL-6 ≥ 3 times the upper limit of normal value

[0462] 5. Females of childbearing potential agree and commit to use an acceptable form of birth control throughout the study. Acceptable forms of birth control in this study are defined as barrier methods plus hormonal therapy (implants, injections, oral contraceptives, and intrauterine devices) or abstinence.

[0463] 6.3.1.1. Exclusion criteria

[0464] Patients who met the following criteria were excluded from the study:

[0465] 1. Patients weighing more than 100 kg

[0466] 2. Alanine aminotransferase / aspartate aminotransferase (ALT / AST)>5 times the upper limit of normal value.

[0467] Stage 3.4 severe chronic kidney disease or requiring dialysis (i.e. estimated glomerular filtration rate (eGFR) <30 ml / min / 1.73 m^2)

[0468] 4. Hemoglobin <80 g / L

[0469] 5. White blood cells <2.0×10^9

[0470] 6. Platelets <50×10^9

[0471] 7. Pregnancy or breast-feeding.

[0472] 8. Expected to be transferred to another hospital other than the research center within 72 hours.

[0473] 9. Life expectancy does not exceed 7 days.

[0474] 10. Patients who have used study medication within 30 days after the first dose of CER-001.

[0475] 6.3.1.2. Restrictions during the study

[0476] There were no patient restrictions other than those outlined in the inclusion / exclusion criteria above.

[0477] 6.3.1.3. Exit criteria

[0478] Reasons for patients to withdraw from study medication may include, but are not limited to, the following:

[0479] · Investigator's request, for safety reasons, such as serious adverse reactions;

[0480] · Investigator's request for other reasons, such as patient non-compliance;

[0481] Patient's request, for tolerability reasons;

[0482] Patient's request, for other reasons, such as withdrawal of informed consent;

[0483] Discontinuation of study drug alone does not constitute discontinuation or withdrawal from the study. Patients will continue to be followed as if they had completed the treatment period. Patients who discontinue study drug prematurely (e.g., before completing the seventh dose) should receive an end-of-study evaluation if possible.

[0484] 6.3.2. Treatment of patients

[0485] 6.3.2.1. Research products

[0486] CER-001 is supplied frozen in 20 mL vials containing approximately 18 mL of product at a concentration of 8 mg / mL (ApoA-I content). CER-001 is dosed by weight. All doses are thawed and then diluted to a volume of 250 mL with normal saline.

[0487] The drug was administered at each of the seven dosing visits. At each visit, patients were given a single IV infusion of CER001 (20 mg / kg) over 24 hours using an infusion pump. Prior to each dose of CER-001, patients were pretreated with an antihistamine (e.g., dexchlorpheniramine 5 mg or hydroxyzine 100 mg) to avoid any potential infusion reactions.

[0488] 6.3.2.2. Interruption or discontinuation of study medication

[0489] Patients interrupted or stopped study drug treatment if any of the following occurred:

[0490] Any drug-related adverse event or event that, in the opinion of the investigator, jeopardizes the patient's participation in the trial or the interpretation of trial data (e.g., a serious intercurrent illness that requires additional nursing care or prevents further dosing)

[0491] Major tolerability issues

[0492] When study medication was discontinued, the study center recorded the reason for discontinuation. Patients were followed up clinically and attempts were made to restart study medication within 2 days of discontinuation if there were no other contraindications.

[0493] 6.3.3. Combined treatment

[0494] All non-experimental treatments were allowed to be taken concurrently during the patient's participation in this study. Any medication taken by the patient in addition to the study medication specified in the protocol was considered as concomitant medication and was recorded in the study records.

[0495] 6.3.4. Prohibited drugs

[0496] There are no excluded medications.

[0497] 6.3.5. Monitoring patient compliance

[0498] CER-001 was administered in a hospital under direct observation.

[0499] 6.3.6. Efficacy evaluation

[0500] 6.3.6.1. Efficacy evaluation

[0501] Inflammatory markers included: CRP, D-dimer, ferritin, IL-6, IL-8, GM-CSF, MCP1, and TNF-α.

[0502] 6.3.6.2. Validity parameters

[0503] (a) Main efficacy parameters

[0504] The primary efficacy parameter was the change in IL-6 from baseline to Day 8. Baseline was defined as the mean of the baseline visit and the Day 1 predose measurement.

[0505] (b) Secondary efficacy parameters

[0506] Secondary efficacy parameters included changes from baseline to day 8 in the inflammatory markers CRP, D-dimer, ferritin, IL-8, GM-CSF, MCP1, and TNF-α.

[0507] 6.3.7. Safety assessment

[0508] 6.3.7.1. Security parameters

[0509] (a) Pregnancy test (if applicable)

[0510] Females of childbearing potential had a documented negative pregnancy test at any time during hospitalization and prior to dosing.

[0511] (b) Safety laboratory testing

[0512] Blood samples were drawn for chemistry and hematology analysis at two time points: baseline and day 8. The following tests were performed by the local hospital laboratory:

[0513]

[0514] Results

[0515] IL-6 levels decreased from baseline to day 8. Secondary efficacy parameters also decreased from baseline to day 8, indicating that CER-001 therapy can be used to treat CRS and reduce serum levels of inflammatory markers.

[0516] 6.4. Example 4: CER-001 Therapy for the Treatment of CRS Secondary to Covid-19 Infection - Additional Treatment Regimens

[0517] This example is a study of CER-001 therapy in COVID-19 patients with severe cytokine release syndrome and renal impairment.

[0518] 6.4.1. Selection of subjects

[0519] 6.4.1.1. Inclusion criteria

[0520] Eligible patients met the following criteria before study enrollment:

[0521] 1. Male or non-pregnant female adults aged 18 years or above at the time of enrollment.

[0522] 2. Have laboratory-confirmed novel coronavirus infection identified by polymerase chain reaction (PCR) or other commercial or public health assays in oropharyngeal or anal specimens within 72 hours prior to hospitalization.

[0523] 3. Illness of any duration and at least one of the following:

[0524] Radiographic infiltration (chest x-ray, CT scan, etc.), or

[0525] Clinical assessment (evidence of rales / crackles on physical examination) and SpO2 < 93% on room air, or requirement for mechanical ventilation and / or supplemental oxygen, or

[0526] Fever that has persisted for the past 24 hours and has not responded to NSAIDs or steroids

[0527] 4. Serum IL-6> 3 times the upper limit of normal

[0528] 5. Females of childbearing potential agree and commit to use an acceptable form of birth control throughout the study. Acceptable forms of birth control in this study are defined as barrier methods plus hormonal therapy (implants, injections, oral contraceptives, and intrauterine devices) or abstinence.

[0529] 6.4.1.2. Exclusion criteria

[0530] Patients who met any of the following criteria were excluded from this study.

[0531] 1. Clinical history suggests allergy to CER-001

[0532] 2. Pregnancy or breastfeeding.

[0533] 3. Expected to be transferred to another hospital within 72 hours.

[0534] 4. Life expectancy does not exceed 7 days.

[0535] 5. Patients who have used study medication within 30 days after the first dose of CER-001.

[0536] 6.4.2. Treatment

[0537] 6.4.2.1. Treatment administered

[0538] Prior to each CER-001 dose, patients were pretreated with an antihistamine (eg, dexchlorpheniramine 5 mg or hydroxyzine 100 mg) to avoid any potential infusion reactions.

[0539] Patients received CER-001 intravenously at a dose of 15 mg / kg BID for 3 consecutive days. Patients could receive up to two additional doses at the investigator's discretion.

[0540] Patients may interrupt or stop study drug treatment if any of the following occur:

[0541] 1. Any drug-related adverse events or other reasons that the investigator believes jeopardize the patient's participation in the trial or the interpretation of trial data (for example, serious complications that require additional care measures or prevent further medication);

[0542] 2. Major tolerance issues.

[0543] When study medication was discontinued, the study center recorded the reason for discontinuation. Patients were followed up clinically and attempts were made to restart study medication within 2 days of discontinuation if there were no other contraindications.

[0544] Reasons for withdrawal from study medication may include, but are not limited to, the following:

[0545] 1. The researcher requires that for safety reasons, such as serious adverse reactions

[0546] 2. The investigator requested that for other reasons, such as patient non-compliance

[0547] 3. Patient request, for tolerability reasons

[0548] 4. Patient’s request, for other reasons, such as withdrawal of informed consent

[0549] Discontinuation of study drug alone does not constitute discontinuation or withdrawal from the study. Patients will continue to be followed as if they had completed the treatment period. Patients who discontinue study drug prematurely (e.g., before completing the third dose) should receive an end-of-study evaluation if possible.

[0550] 6.4.2.2. Dose changes

[0551] The dose of the drug may be reduced or increased if clinically necessary as determined by the principal investigator.

[0552] 6.4.2.3. Concomitant medication / therapy

[0553] All concomitant non-experimental treatments were permitted during the patient's participation in this study. Any medication taken by the patient in addition to the study medication specified in the protocol was considered concomitant medication and was recorded in the study records.

[0554] 6.4.3. Research evaluation

[0555] The following procedures were performed during the baseline visit. The following tests were performed by the local hospital laboratory.

[0556] 1. Informed Consent

[0557] 2. Medical history included recording of past and present illnesses and collection of subject demographic data (date of birth, sex, and race).

[0558] 3. Physical examination, including review of systems, height and weight, BMI, and waist circumference

[0559] 4. Vital signs (pulse, blood pressure, and oral, aural, axillary, or core temperature).

[0560] 5. Review of inclusion / exclusion criteria.

[0561] 6. Record adverse events starting from the time of obtaining informed consent.

[0562] 7. Collect the previous medications taken 4 weeks before the first dose of the test article. Record all current medications.

[0563] 8. Complete blood count (CBC) - includes white blood cell count (WBC) and differential, platelet count, red blood cell count (RBC), hemoglobin (Hb), hematocrit (Hct).

[0564] 9. Fasting chemistry panel / electrolytes: including sodium, potassium, chloride, blood urea nitrogen (BUN; or urea), serum creatinine, calculated creatinine clearance (CKD-EPI), glucose, calcium, phosphorus, total protein, uric acid, AST, ALT, γGT, ALP, total and direct bilirubin, albumin, total cholesterol, HDL, LDL, triglycerides, LDH, CPK,

[0565] 10. ABG (to evaluate respiratory and / or metabolic disorders)

[0566] 11. ApoA-I (for pharmacokinetic and pharmacodynamic evaluation)

[0567] 12. Coagulation tests - including prothrombin time (PT) (expressed as international normalized ratio [INR]) and partial thromboplastin time (PTT).

[0568] 13. Urinalysis - includes assessment of specific gravity, pH, protein / albumin, glucose, ketones, and hemoglobin / blood.

[0569] 14. Microalbuminuria and proteinuria g / 24h

[0570] 15. Serum or urine pregnancy test within 7 days before randomization (for women of childbearing potential).

[0571] 16. Pharmacokinetic and pharmacodynamic evaluations including apoA-I and total cholesterol levels.

[0572] 17. Inflammatory markers include CRP, PCT, D-dimer, ferritin, IL-6, IL-8, GM-CSF, MCP1 and TNF-α.

[0573] like Figure 7 Clinical and laboratory parameters were monitored from baseline to the last visit on Day 8, as reported, and included the following procedures:

[0574] 1. Record adverse events and concomitant medications

[0575] 2. Review appropriate laboratory information

[0576] 3. Physical examination

[0577] 4. Assess vital signs (pulse, blood pressure, and oral, aural, axillary, or core temperature)

[0578] 5. Keep records of adverse events and concomitant medications

[0579] 6. Complete blood count (CBC) - includes white blood cell count (WBC) and differential, platelet count, red blood cell count (RBC), hemoglobin (Hb), hematocrit (Hct).

[0580] 7. Fasting chemistry panel / electrolytes: including sodium, potassium, chloride, blood urea nitrogen (BUN; or urea), serum creatinine, calculated creatinine clearance (CKD-EPI), glucose, calcium, phosphorus, total protein, uric acid, AST, ALT, γGT, ALP, total and direct bilirubin, albumin, total cholesterol, HDL, LDL, triglycerides, LDH, CPK

[0581] 8. ABG (to evaluate respiratory and / or metabolic disturbances)

[0582] 9. ApoA-I (for pharmacokinetic and pharmacodynamic evaluation)

[0583] 10. Coagulation tests - including prothrombin time (PT) (expressed as international normalized ratio [INR]) and partial thromboplastin time (PTT).

[0584] 11. Urinalysis - includes assessment of specific gravity, pH, protein / albumin, glucose, ketones, and hemoglobin / blood.

[0585] 12. Microalbuminuria and proteinuria g / 24h

[0586] 13. Inflammatory markers include CRP, PCT, D-dimer, ferritin, IL-6, IL-8, GM-CSF, MCP1 and TNF-α

[0587] 6.4.4. Adverse Event (AE) Reporting

[0588] An AE is any adverse medical event associated with the use of an investigational product (active or placebo drug, biologic, or device) in a clinical study patient that does not necessarily have a causal relationship to the product. Thus, an AE can be any unfavorable and unexpected sign (e.g., abnormal laboratory test results), symptom, or disease temporally associated with the use of an investigational product, whether or not considered related to the investigational product.

[0589] Adverse events may include:

[0590] Symptoms described by the patient

[0591] Clinically significant changes in the patient's physical examination or other signs observed by the investigator or medical staff

[0592] Abnormal tests (laboratory tests) that reflect a change from baseline and / or that may lead to a change in the administration of the investigational product or a change in medical care (diagnostic or therapeutic)

[0593] A condition that was present at baseline but then resolved and then worsened or recurred

[0594] At each study visit, patients were assessed for the status of new and existing AEs.

[0595] Results

[0596] IL-6 levels and other inflammatory markers decreased from baseline to day 8.

[0597] 6.5. Example 5: CER-001 Therapy for the Treatment of Ischemia / Reperfusion AKI

[0598] This example is a study of CER-001 therapy for the treatment of ischemia / reperfusion AKI.

[0599] Materials and methods

[0600] Pigs weighing 45-60kg were fasted for 24 hours before the study. All animals were pre-administered with an intramuscular mixture of azaperidone (8mg kg-1) and atropine (0.03mg kg-1) to reduce pharyngeal and tracheal secretions and prevent bradycardia after intubation. After anesthesia, the kidneys were approached through a midline abdominal incision. Then, the renal artery and vein were separated, and the vascular ring was positioned around the renal artery with a right-angle clamp. Warm ischemia was induced by pulling the vascular ring for 60 minutes. Ischemia was followed by 3 hours of reperfusion, half of which received CER-001 directly administered through the renal artery 5 minutes before the start of reperfusion. The animals were euthanized by IV administration of 1mL / kg BW pentobarbital 24 hours later. The kidneys were then harvested for analysis.

[0601] Results

[0602] CER-001 alleviates ischemia / reperfusion AKI.

[0603] 6.6. Example 6: Lipid Binding Protein Molecular Therapy in LPS-Induced Endothelial Injury Model

[0604] The ability of the ApoA-I-containing complex CER-001 to mitigate sepsis-induced endothelial injury was evaluated in an in vitro model induced by lipopolysaccharide (LPS).

[0605] Materials and methods

[0606] 6.6.1.1. Cell culture

[0607] Human umbilical vein endothelial cells (HUVEC, EC) were purchased from the American Type Culture Collection (ATCC-LGCStandards Srl, Sesto San Giovanni, Milan, Italy). EC were maintained in EndGro, its recommended medium (Merck Millipore, Darmstadt, Germany).

[0608] As previously described (Sallustio, et al., 2021, Nephrol Dial Transplant 36, 452-464), peripheral blood mononuclear cells (PBMCs) were isolated from the buffy coat of healthy donors (selected from the research bank) by gradient centrifugation using the Ficoll-Hypaque method. PBMCs were maintained in their recommended culture medium (supra).

[0609] When cells were confluent, they were stimulated with LPS 0.3 μg / ml, 4 μg / ml (E. coli O111:B4, Sigma-Aldrich, Milan, Italy) and CER-001 50 μg / ml, 100 μg / ml and 500 μg / ml for the indicated periods of time.

[0610] PBMC culture supernatants were collected and analyzed for TNF-α by ELISA (R&D Systems, Minneapolis MN, USA).

[0611] 6.6.1.2. Cell proliferation assay

[0612] EC and PBMC were incubated with 0.3 μg / ml LPS and / or 50 μg / ml and 500 μg / ml CER-001 for 60 min and 24 h. Proliferation rate was measured by MTT cell proliferation assay kit according to the manufacturer's instructions (Sigma Aldrich). 4 Cells / well were seeded in 96-well plates, and then the cells were treated with LPS and CER-001 as indicated. The absorbance at 570 nm was then measured by a spectrophotometer.

[0613] 6.6.1.3. Immunophenotyping

[0614] After stimulation, ECs were permeabilized using the IntraPrep kit (Instrumentation Laboratory) and incubated with unconjugated primary antibody p-ENOS (Abcam) for 25 min at 4° C. Cells were then washed and labeled with secondary antibody AlexaFluor 488 (Molecular Probes) for 25 min at 4° C. Finally, cells were washed twice and resuspended in FACS buffer for acquisition.

[0615] PBMCs were stained with the following monoclonal antibody CD14 monoclonal antibody (61D3)-PE (eBioscience Italy, Thermo Fisher Scientific, Italy) at room temperature in the dark for 20 minutes, washed twice, and resuspended in FACS buffer. The stained PBMCs were then obtained.

[0616] Data were obtained by using a FC500 (Beckman Coulter) flow cytometer and analyzed with Kaluza software. Three independent studies were performed on EC and PBMC. Positive areas were determined by using isotype-matched mAbs, and a total of 104 events were obtained for each sample.

[0617] Statistical analysis

[0618] The data shown are representative of three independent studies. Data are shown as mean ± standard deviation (SD) and compared using Student-t test.

[0619] Results

[0620] The effects of LPS and CER-001 on endothelial cells and endothelial nitric oxide synthase (eNOS) activation were analyzed. Fig.11 The results of the MTT cell viability assay in the Figure 3 showed a slight decrease in proliferation after LPS stimulation. CER-001 at 50 μg / ml and 500 μg / ml did not affect endothelial viability. Endothelial cells treated with LPS and CER-001 at 50 μg / ml and 500 μg / ml increased the proliferation rate compared to LPS-stimulated cells, especially at the highest concentration.

[0621] The production of eNOS has been described as a marker of endothelial integrity (Zhao, et al., 2015, J Pharmacol Sci 129, 83-94). In the in vitro model of this example, eNOS phosphorylation and activation ( Figure 12-13 ) was altered by LPS and upregulated by CER-001. Specifically, a strong decrease in eNOS (phospho-S1177) (p-ENOS) was observed after 60 min of LPS stimulation compared to basal and VEGF (positive control). Supplementation with 500 μg / mL CER-001 completely reversed the LPS effect. (In Fig.13 Representative data from one of a total of three studies are shown. Histograms represent p-ENOS expression levels).

[0622] Furthermore, CER-001 modulated the responses of peripheral blood mononuclear cells (PBMCs) stimulated with 0.3 μg / mL LPS and / or 50 μg / mL and 500 μg / mL CER-001 for 24 hours, reducing mCD14 expression and TNF-α secretion. Fig.14 As shown, for the above conditions, the MTT assay showed no significant difference in cell viability relative to the basal level. The PBMC culture supernatants were analyzed by ELISA, and the results were shown in Fig.15 As shown. PBMC increased TNF-α synthesis 24 hours after LPS stimulation. Stimulation of PBMC with 50μg / m and 500μg / m CER-001 alone did not affect TNF-α production. Addition of 50μg / m and 500μg / m CER-001 to the culture medium of LPS-activated PBMC reversed the LPS effect. In addition, FACS showed a strong upregulation of CD14 surface expression by PCMB 24 hours after LPS stimulation ( Fig.16PBMCs treated with a combination of LPS and CER-001 maintained CD14 expression at basal levels ( Fig.17 ).

[0623] 7. Incorporation by Reference

[0624] All publications, patents, patent applications, and other documents cited in this application are incorporated herein by reference in their entirety for all purposes to the same extent as if each individual publication, patent, patent application, or other document was individually indicated to be incorporated by reference for all purposes.

[0625] Any discussion of documents, acts, materials, devices, articles of manufacture or the like that has been included in this specification is solely for the purpose of providing a context for the present disclosure. This should not be taken as an admission that any or all of these matters form part of the prior art base or are common general knowledge in the field relevant to the present disclosure as it existed anywhere before the priority date of this application. DETAILED DESCRIPTION

[0627] Various aspects of the present disclosure are described in the embodiments set forth in the following numbered paragraphs of Group 1.

[0628] Group 1:

[0629] 1. A method of treating a subject having or at risk of developing leukocytosis, the method comprising administering to the subject a lipid binding protein-based complex in an amount effective to reduce the subject's leukocyte count.

[0630] 2. The method of embodiment 1, wherein the subject suffers from one or more symptoms associated with leukocytosis.

[0631] 3. A method of treating a subject suffering from one or more symptoms associated with leukocytosis, the method comprising administering to the subject an amount of a lipid binding protein-based complex effective to reduce the subject's white blood cell count and / or improve one or more of the one or more symptoms associated with leukocytosis.

[0632] 4. The method of embodiment 2 or embodiment 3, wherein the one or more symptoms include fever.

[0633] 5. The method of any one of embodiments 2 to 4, wherein the one or more symptoms include bleeding or bruising.

[0634] 6. The method of any one of embodiments 2 to 5, wherein the one or more symptoms include sweating.

[0635] 7. The method of any one of embodiments 2 to 6, wherein the one or more symptoms comprise pain or tingling in the legs, arms, or abdomen.

[0636] 8. The method of any one of embodiments 2 to 7, wherein the one or more symptoms include one or more vision problems.

[0637] 9. The method of embodiment 8, wherein the one or more vision problems include blurred vision.

[0638] 10. The method of embodiment 8 or embodiment 9, wherein the one or more vision problems include diplopia.

[0639] 11. The method of any one of embodiments 8 to 10, wherein the one or more vision problems include a blind spot.

[0640] 12. The method of any one of embodiments 8 to 11, wherein the one or more vision problems include blurred vision.

[0641] 13. The method of any one of embodiments 8 to 12, wherein the one or more vision problems comprise blurred vision.

[0642] 14. The method of any one of embodiments 2 to 13, wherein the one or more symptoms include unclear thinking.

[0643] 15. The method of any one of embodiments 2 to 14, wherein the one or more symptoms include loss of appetite.

[0644] 16. The method of any one of embodiments 2 to 15, wherein the one or more symptoms include difficulty breathing (e.g., shortness of breath, below normal blood oxygen levels).

[0645] 17. The method of any one of embodiments 2 to 16, wherein the dose of the lipid binding protein-based complex is effective to improve one or more of the one or more symptoms associated with leukocytosis.

[0646] 18. The method of any one of embodiments 1 to 17, wherein the subject has or is at risk of developing leukocytosis due to inflammation, infection, a leukocyte disorder, physical stress, emotional stress, a medication, or an allergic reaction.

[0647] 19. The method of embodiment 18, wherein the subject has or is at risk of developing leukocytosis due to inflammation.

[0648] 20. The method of embodiment 18, wherein the subject has or is at risk of developing leukocytosis due to an infection.

[0649] 21. The method of embodiment 18, wherein the subject has or is at risk of developing leukocytosis due to a leukocyte disorder.

[0650] 22. The method of embodiment 18, wherein the subject suffers from or is at risk of developing leukocytosis due to physical stress, optionally wherein the physical stress is from an epileptic seizure, anesthesia, or overexertion.

[0651] 23. The method of embodiment 18, wherein the subject suffers from or is at risk of developing leukocytosis due to emotional stress.

[0652] 24. The method of embodiment 18, wherein the subject has or is at risk of developing leukocytosis due to a medication, optionally wherein the medication is a steroid, a corticosteroid, lithium, or a beta agonist.

[0653] 25. The method of embodiment 18, wherein the subject has or is at risk of developing leukocytosis due to an allergic reaction.

[0654] 26. The method of any one of embodiments 1 to 25, wherein the subject has an infection.

[0655] 27. The method of embodiment 26, wherein the infection is a viral infection, optionally wherein the viral infection is a coronavirus infection, which is optionally COVID-19.

[0656] 28. The method of embodiment 26, wherein the infection is a bacterial infection.

[0657] 29. The method of embodiment 26, wherein the infection is a fungal infection.

[0658] 30. The method of embodiment 26, wherein the infection is a parasitic infection.

[0659] 31. The method of any one of embodiments 1 to 30, wherein the subject has diabetes.

[0660] 32. The method of any one of embodiments 1 to 31, wherein the subject has a leukocyte disorder.

[0661] 33. The method of embodiment 32, wherein the leukocyte disorder is a primary bone marrow disorder (eg, an acute leukemia, a chronic leukemia, or a myeloproliferative disorder).

[0662] 34. The method of embodiment 32, wherein the leukocyte disorder is neutrophilia, optionally wherein the neutrophilia is idiopathic neutrophilia, which is optionally chronic idiopathic neutrophilia.

[0663] 35. The method of embodiment 32, wherein the leukocyte disorder is hemolytic anemia.

[0664] 36. The method of embodiment 32, wherein the leukocyte disorder is thrombocytopenia, optionally wherein the thrombocytopenia is idiopathic thrombocytopenia or essential thrombocytopenia.

[0665] 37. The method of embodiment 32, wherein the leukocyte disorder is cancer, optionally bone cancer or blood cancer.

[0666] 38. The method of embodiment 32, wherein the leukocyte disorder is a myeloproliferative disorder or a blood cancer.

[0667] 39. The method of embodiment 38, wherein the leukocyte disorder is a myeloproliferative disorder.

[0668] 40. The method of embodiment 38, wherein the leukocyte disorder is a blood cancer.

[0669] 41. The method of embodiment 38, wherein the leukocyte disorder is lymphoma.

[0670] 42. The method of embodiment 38, wherein the white blood cell disorder is leukemia.

[0671] 43. The method of embodiment 42, wherein the leukemia is chronic leukemia.

[0672] 44. The method of embodiment 42, wherein the leukemia is acute leukemia.

[0673] 45. The method of embodiment 42, wherein the leukemia is lymphocytic leukemia, chronic eosinophilic leukemia, chronic myeloid leukemia, or chronic neutrophilic leukemia.

[0674] 46. ​​The method of embodiment 38, wherein the leukocyte disorder is polycythemia vera.

[0675] 47. The method of embodiment 38, wherein the leukocyte disorder is myelofibrosis, optionally wherein the myelofibrosis is primary myelofibrosis.

[0676] 48. The method of any one of embodiments 1 to 47, wherein the subject has leukocytosis.

[0677] 49. The method of embodiment 48, wherein the subject has a serum serotype higher than 11 x 10 9 / L white blood cell count.

[0678] 50. The method of embodiment 48 or embodiment 49, wherein the subject has a serum serotype greater than 12 x 10 9 / L white blood cell count.

[0679] 51. The method of any one of embodiments 48 to 50, wherein the subject has a serum serotype greater than 15 x 10 9 / L white blood cell count.

[0680] 52. The method of any one of embodiments 48 to 51, wherein the subject has a serum serotype greater than 20 x 10 9 / L white blood cell count.

[0681] 53. A method as described in embodiment 48 or embodiment 49, wherein the subject has less than 100x10 9 / L white blood cell count.

[0682] 54. The method of any one of embodiments 48 to 52, wherein the subject has less than 100 x 10 9 / L white blood cell count is lower than.

[0683] 55. The method of any one of embodiments 48 to 54, wherein the subject has less than 50 x 10 9 / L white blood cell count.

[0684] 56. The method of any one of embodiments 48 to 55, wherein the subject has less than 25 x 10 9 / L white blood cell count.

[0685] 57. The method of any one of embodiments 48 to 56, wherein the subject has less than 15 x 10 9 / L white blood cell count.

[0686] 58. The method of any one of embodiments 48 to 57, wherein the subject has a 9 / L and 50x10 9 / L between the white blood cell count.

[0687] 59. The method of any one of embodiments 48 to 57, wherein the subject has a 9 / L and 25x10 9 / L between the white blood cell count.

[0688] 60. The method of any one of embodiments 48 to 57, wherein the subject has a 9 / L and 50x10 9 / L between the white blood cell count.

[0689] 61. The method of any one of embodiments 48 to 51, wherein the subject has a 9 / L and 100x10 9 / L between the white blood cell count.

[0690] 62. The method of any one of embodiments 48 to 57, wherein the subject has a 9 / L and 15x10 9 / L between the white blood cell count.

[0691] 63. The method of any one of embodiments 48 to 57, wherein the subject has a 9 / L and 20x10 9 / L between the white blood cell count.

[0692] 64. The method of any one of embodiments 1 to 47, wherein the subject is at risk for leukocytosis.

[0693] 65. A method of treating a subject suffering from endothelial dysfunction, the method comprising administering to the subject a dose of a lipid binding protein-based complex, wherein the dose is a high dose.

[0694] 66. A method of treating a subject who is experiencing or has experienced an acute coronary syndrome or stroke, the method comprising administering to the subject a dose of a lipid binding protein-based complex, wherein the dose is a high dose.

[0695] 67. The method of embodiment 65 or embodiment 66, wherein the subject is experiencing an acute coronary syndrome, optionally wherein the acute coronary syndrome is myocardial infarction (e.g., ST-elevation myocardial infarction or non-ST-elevation myocardial infarction) or unstable angina.

[0696] 68. The method of embodiment 65 or embodiment 66, wherein the subject has experienced an acute coronary syndrome, optionally wherein the acute coronary syndrome is myocardial infarction (e.g., ST-elevation myocardial infarction or non-ST-elevation myocardial infarction) or unstable angina.

[0697] 69. The method of embodiment 65 or embodiment 66, wherein the subject is experiencing a stroke.

[0698] 70. The method of embodiment 65 or embodiment 66, wherein the subject has experienced a stroke.

[0699] 71. A method of treating a subject having or at risk of developing carditis, the method comprising administering to the subject a dose of a lipid binding protein-based complex, optionally wherein the carditis is myocarditis and / or pericarditis.

[0700] 72. The method of embodiment 71, wherein the carditis comprises myocarditis.

[0701] 73. The method of embodiment 71 or embodiment 72, wherein the carditis comprises pericarditis.

[0702] 74. The method of any one of embodiments 71 to 73, wherein the subject is male.

[0703] 75. The method of any one of embodiments 71 to 74, wherein the subject is less than 60 years old.

[0704] 76. The method of any one of embodiments 71 to 74, wherein the subject is less than 50 years old.

[0705] 77. The method of any one of embodiments 71 to 74, wherein the subject is less than 40 years old.

[0706] 78. The method of any one of embodiments 71 to 74, wherein the subject is less than 30 years old.

[0707] 79. The method of any one of embodiments 71 to 74, wherein the subject is less than 20 years old.

[0708] 80. The method of any one of embodiments 71 to 79, wherein the subject is at least 5 years old.

[0709] 81. The method of any one of embodiments 71 to 79, wherein the subject is at least 8 years old.

[0710] 82. The method of any one of embodiments 71 to 79, wherein the subject is at least 10 years old.

[0711] 83. The method of any one of embodiments 71 to 79, wherein the subject is at least 12 years old.

[0712] 84. The method of any one of embodiments 71 to 79, wherein the subject is at least 15 years old.

[0713] 85. The method of any one of embodiments 71 to 79, wherein the subject is at least 18 years old.

[0714] 86. The method of any one of embodiments 71 to 85, wherein the subject has an infection.

[0715] 87. The method of embodiment 74, wherein the infection is a viral infection, optionally a coronavirus infection, optionally COVID-19.

[0716] 88. The method of embodiment 74, wherein the infection is a bacterial infection.

[0717] 89. The method of any one of embodiments 71 to 88, wherein the lipid binding protein-based complex is administered prior to administering a vaccine to the subject.

[0718] 90. The method of any one of embodiments 71 to 88, wherein the lipid binding protein-based complex is administered to the subject simultaneously with a vaccine.

[0719] 91. The method of any one of embodiments 71 to 88, wherein the lipid binding protein-based complex is administered after administration of a vaccine to the subject.

[0720] 92. The method of any one of embodiments 89 to 91, wherein the vaccine is a coronavirus vaccine, optionally a COVID-19 vaccine.

[0721] 93. The method of any one of embodiments 71 to 92, wherein the subject has carditis.

[0722] 94. The method of embodiment 93, wherein the dose of the lipid binding protein-based complex is effective to improve one or more symptoms of the carditis.

[0723] 95. The method of any one of embodiments 71 to 92, wherein the subject is at risk for carditis.

[0724] 96. The method of embodiment 95, wherein the dose of the lipid binding protein-based complex is effective to prevent carditis or reduce the severity of carditis.

[0725] 97. The method of any one of embodiments 1 to 96, wherein the dose is effective to reduce circulating VCAM-1 and / or ICAM-1 in the subject.

[0726] 98. The method of embodiment 97, wherein the dose is effective to reduce circulating VCAM-1 in the subject by at least 100 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0727] 99. The method of embodiment 97, wherein the dose is effective to reduce the subject's circulating VCAM-1 by at least 200 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0728] 100. The method of embodiment 97, wherein the dose is effective to reduce the subject's circulating VCAM-1 by at least 300 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0729] 101. The method of any one of embodiments 97 to 100, wherein the dose is effective to reduce the subject's circulating VCAM-1 by up to 400 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0730] 102. The method of embodiments 97 to 101, wherein the dose is effective to reduce circulating VCAM-1 in the subject by at least 100 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0731] 103. The method of embodiments 97 to 101, wherein the dose is effective to reduce the subject's circulating VCAM-1 by at least 200 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0732] 104. The method of embodiments 97 to 101, wherein the dose is effective to reduce the subject's circulating VCAM-1 by at least 300 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0733] 105. The method of any one of embodiments 97 to 104, wherein the dose is effective to reduce the subject's circulating VCAM-1 by up to 400 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0734] 106. The method of any one of embodiments 97 to 105, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 50 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0735] 107. The method of any one of embodiments 97 to 105, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 75 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0736] 108. The method of any one of embodiments 97 to 105, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 100 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0737] 109. The method of any one of embodiments 97 to 108, wherein the dose is effective to reduce the subject's circulating ICAM-1 by up to 125 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0738] 110. The method of any one of embodiments 97 to 110, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 50 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0739] 111. The method of any one of embodiments 97 to 110, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 75 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0740] 112. The method of any one of embodiments 97 to 110, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 100 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0741] 113. The method of any one of embodiments 97 to 112, wherein the dose is effective to reduce the subject's circulating ICAM-1 by up to 125 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0742] 114. The method of any one of embodiments 1 to 113, wherein the dose is a high dose.

[0743] 115. The method of embodiment 114, wherein the high dose is administered over a period of two days to about two weeks, optionally wherein the high dose is administered over a period of two, three, four, five, six, seven, eight, nine, 10, 11, 12, 13, 14, or 15 days.

[0744] 116. The method of embodiment 115, wherein the high dose is administered over a period of two days.

[0745] 117. The method of embodiment 115, wherein the high dose is administered over a period of three days.

[0746] 118. The method of embodiment 115, wherein the high dose is administered over a four-day period.

[0747] 119. The method of embodiment 115, wherein said dose is administered over a period of five days.

[0748] 120. The method of embodiment 115, wherein the high dose is administered over a period of six days.

[0749] 121. The method of embodiment 115, wherein the high dose is administered over a period of seven days.

[0750] 122. The method of embodiment 115, wherein the high dose is administered over a period of eight days.

[0751] 123. The method of embodiment 115, wherein the high dose is administered over a period of nine days.

[0752] 124. The method of embodiment 115, wherein the high dose is administered over a period of 10 days.

[0753] 125. The method of embodiment 115, wherein the high dose is administered over a period of 11 days.

[0754] 126. The method of embodiment 115, wherein the high dose is administered over a period of 12 days.

[0755] 127. The method of embodiment 115, wherein the high dose is administered over a period of 13 days.

[0756] 128. The method of embodiment 115, wherein the high dose is administered over a period of 14 days.

[0757] 129. The method of embodiment 115, wherein the high dose is administered over a period of 15 days.

[0758] 130. The method of any one of embodiments 114 to 129, wherein the high dose is the sum of two to 20 individual doses, optionally wherein the high dose is the sum of two, three, four, five, six, seven, eight, nine or 10 individual doses.

[0759] 131. The method of embodiment 130, wherein the high dose comprises two to 16 separate doses.

[0760] 132. The method of embodiment 130, wherein the high dose comprises two to 12 separate doses.

[0761] 133. The method of embodiment 130, wherein the high dose comprises two to 10 separate doses.

[0762] 134. The method of embodiment 130, wherein the high dose comprises two to eight separate doses.

[0763] 135. The method of embodiment 130, wherein the high dose comprises two to six separate doses.

[0764] 136. The method of embodiment 130, wherein the high dose comprises two to four separate doses.

[0765] 137. The method of embodiment 130, wherein the high dose comprises four to 20 separate doses.

[0766] 138. The method of embodiment 130, wherein the high dose comprises four to 16 separate doses.

[0767] 139. The method of embodiment 130, wherein the high dose comprises four to ten separate doses.

[0768] 140. The method of embodiment 130, wherein the high dose comprises four to six separate doses.

[0769] 141. The method of embodiment 130, wherein the high dose comprises six to 20 individual doses.

[0770] 142. The method of embodiment 130, wherein the high dose comprises six to 16 separate doses.

[0771] 143. The method of embodiment 130, wherein the high dose comprises six to ten separate doses.

[0772] 144. The method of embodiment 130, wherein the high dose comprises six to eight individual doses.

[0773] 145. The method of embodiment 130, wherein the high dose comprises eight to 20 individual doses.

[0774] 146. The method of embodiment 130, wherein the high dose comprises eight to 16 separate doses.

[0775] 147. The method of embodiment 130, wherein the high dose comprises eight to ten separate doses.

[0776] 148. The method of embodiment 130, wherein the high dose comprises 10 to 20 individual doses.

[0777] 149. The method of embodiment 130, wherein the high dose comprises 10 to 16 individual doses.

[0778] 150. The method of embodiment 130, wherein the high dose comprises 16 to 20 individual doses.

[0779] 151. The method of embodiment 130, wherein the high dose comprises two separate doses.

[0780] 152. The method of embodiment 130, wherein the high dose comprises three separate doses.

[0781] 153. The method of embodiment 130, wherein the high dose comprises four separate doses.

[0782] 154. The method of embodiment 130, wherein the high dose comprises five separate doses.

[0783] 155. The method of embodiment 130, wherein the high dose comprises six separate doses.

[0784] 156. The method of embodiment 130, wherein the high dose comprises seven separate doses.

[0785] 157. The method of embodiment 130, wherein the high dose comprises eight separate doses.

[0786] 158. The method of embodiment 130, wherein the high dose comprises nine separate doses.

[0787] 159. The method of embodiment 130, wherein the high dose comprises ten separate doses.

[0788] 160. The method of any one of embodiments 130 to 159, wherein multiple individual doses are administered daily or twice daily.

[0789] 161. The method of any one of embodiments 130 to 159, wherein multiple individual doses are administered about 12 hours apart.

[0790] 162. The method of any one of embodiments 130 to 161, wherein multiple individual doses are administered two to three days apart.

[0791] 163. The method of any one of embodiments 130 to 162, wherein each individual dose is effective to increase the subject's HDL level.

[0792] 164. The method of embodiment 163, wherein each individual dose is effective to increase the subject's HDL level by at least 25%, at least 30%, or at least 35% 2-4 hours after administration.

[0793] 165. The method of embodiment 164, wherein each individual dose is effective to increase the subject's HDL level by at least 25%, at least 30%, or at least 35% 2 hours after administration.

[0794] 166. The method of embodiment 164, wherein each individual dose is effective to increase the subject's HDL level by at least 25%, at least 30%, or at least 35% 3 hours after administration.

[0795] 167. The method of embodiment 164, wherein each individual dose is effective to increase the subject's HDL level by at least 25%, at least 30%, or at least 35% 4 hours after administration.

[0796] 168. The method of any one of embodiments 130 to 167, wherein each individual dose is effective to increase the subject's ApoA-I level.

[0797] 169. The method of embodiment 168, wherein each individual dose is effective to increase the subject's ApoA-I level by at least 25%, at least 30%, or at least 35% 2-4 hours after administration.

[0798] 170. The method of embodiment 169, wherein each individual dose is effective to increase the subject's ApoA-I level by at least 25%, at least 30%, or at least 35% 2 hours after administration.

[0799] 171. The method of embodiment 169, wherein each individual dose is effective to increase the subject's ApoA-I level by at least 25%, at least 30%, or at least 35% 3 hours after administration.

[0800] 172. The method of embodiment 169, wherein each individual dose is effective to increase the subject's ApoA-I level by at least 25%, at least 30%, or at least 35% 4 hours after administration.

[0801] 173. The method of any one of embodiments 1 to 172, wherein the dose is effective to improve vascular endothelial function in the subject, optionally wherein vascular endothelial function is measured by circulating VCAM-1 and / or ICAM-1.

[0802] 174. The method of embodiment 173, wherein the dose is effective to reduce the subject's circulating VCAM-1 by at least 100 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0803] 175. The method of embodiment 173, wherein the dose is effective to reduce the subject's circulating VCAM-1 by at least 200 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0804] 176. The method of embodiment 173, wherein the dose is effective to reduce the subject's circulating VCAM-1 by at least 300 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0805] 177. The method of any one of embodiments 173 to 176, wherein the dose is effective to reduce the subject's circulating VCAM-1 by up to 400 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0806] 178. The method of embodiments 173 to 177, wherein the dose is effective to reduce circulating VCAM-1 in the subject by at least 100 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0807] 179. The method of embodiments 173 to 177, wherein the dose is effective to reduce the subject's circulating VCAM-1 by at least 200 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0808] 180. The method of embodiments 173 to 177, wherein the dose is effective to reduce circulating VCAM-1 in the subject by at least 300 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0809] 181. The method of any one of embodiments 173 to 180, wherein the dose is effective to reduce the subject's circulating VCAM-1 by up to 400 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0810] 182. The method of any one of embodiments 173 to 181, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 50 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0811] 183. The method of any one of embodiments 173 to 181, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 75 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0812] 184. The method of any one of embodiments 173 to 181, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 100 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0813] 185. The method of any one of embodiments 173 to 184, wherein the dose is effective to reduce the subject's circulating ICAM-1 by up to 125 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0814] 186. The method of any one of embodiments 173 to 185, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 50 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0815] 187. The method of any one of embodiments 173 to 185, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 75 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0816] 188. The method of any one of embodiments 173 to 185, wherein the dose is effective to reduce the subject's circulating ICAM-1 by at least 100 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0817] 189. The method of any one of embodiments 173 to 188, wherein the dose is effective to reduce the subject's circulating ICAM-1 by up to 125 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0818] 190. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 2000 WBC / microliter within two days of the first administration of the lipid binding protein-based complex.

[0819] 191. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 3000 WBC / microliter within two days of the first administration of the lipid binding protein-based complex.

[0820] 192. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 4000 WBC / microliter within two days of the first administration of the lipid binding protein-based complex.

[0821] 193. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 5000 WBC / microliter within two days of the first administration of the lipid binding protein-based complex.

[0822] 194. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 6000 WBC / microliter within two days of the first administration of the lipid binding protein-based complex.

[0823] 195. The method of any one of embodiments 1 to 194, wherein the dose is effective to reduce the subject's white blood cell count by up to 8000 WBC / microliter within two days of the first administration of the lipid binding protein-based complex.

[0824] 196. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 2000 WBC / microliter within five days of the first administration of the lipid binding protein-based complex.

[0825] 197. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 3000 WBC / microliter within five days of the first administration of the lipid binding protein-based complex.

[0826] 198. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 4000 WBC / microliter within five days of the first administration of the lipid binding protein-based complex.

[0827] 199. The method of any one of embodiments 1 to 189, wherein the dose is effective to reduce the subject's white blood cell count by at least 5000 WBC / microliter within five days of the first administration of the lipid binding protein-based complex.

[0828] 200. The method of any one of embodiments 196 to 199, wherein the dose is effective to reduce the subject's white blood cell count by up to 6000 WBC / μL within two days of the first administration of the lipid binding protein-based complex.

[0829] 201. The method of any one of embodiments 1 to 200, wherein the dose is effective to reduce serum levels of one or more inflammatory markers in the subject.

[0830] 202. The method of embodiment 201, wherein the dose is effective to reduce serum levels of interleukin-6 ("IL-6").

[0831] 203. The method of embodiment 202, wherein the dose is effective to reduce the serum level of IL-6 by at least 20 pg / mL within two days of the first administration of the lipid binding protein-based complex.

[0832] 204. The method of embodiment 202, wherein the dose is effective to reduce the serum level of IL-6 by at least 30 pg / mL within two days of the first administration of the lipid binding protein-based complex.

[0833] 205. The method of any one of embodiments 202 to 204, wherein the dose is effective to reduce serum levels of IL-6 by up to 40 pg / mL within two days of the first administration of the lipid binding protein-based complex.

[0834] 206. The method of any one of embodiments 202 to 205, wherein the dose is effective to reduce serum levels of IL-6 by at least 20 pg / mL within five days of the first administration of the lipid binding protein-based complex.

[0835] 207. The method of any one of embodiments 202 to 205, wherein the dose is effective to reduce serum levels of IL-6 by at least 30 pg / mL within five days of the first administration of the lipid binding protein-based complex.

[0836] 208. The method of any one of embodiments 202 to 205, wherein the dose is effective to reduce serum levels of IL-6 by at least 40 pg / mL within five days of the first administration of the lipid binding protein-based complex.

[0837] 209. The method of any one of embodiments 202 to 208, wherein the dose is effective to reduce serum levels of IL-6 by up to 60 pg / mL within five days of the first administration of the lipid binding protein-based complex.

[0838] 210. The method of any one of embodiments 202 to 208, wherein the dose is effective to reduce serum levels of IL-6 by up to 50 pg / mL within five days of the first administration of the lipid binding protein-based complex.

[0839] 211. The method of any one of embodiments 201 to 210, wherein the dose is effective to reduce serum levels of C-reactive protein.

[0840] 212. The method of any one of embodiments 201 to 211, wherein the dose is effective to reduce serum levels of D-dimer.

[0841] 213. The method of any one of embodiments 201 to 212, wherein the dose is effective to reduce serum levels of ferritin.

[0842] 214. The method of embodiment 213, wherein the dose is effective to reduce serum levels of ferritin by at least 200 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0843] 215. The method of embodiment 213, wherein the dose is effective to reduce serum levels of ferritin by at least 300 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0844] 216. The method of embodiment 213, wherein the dose is effective to reduce serum levels of ferritin by at least 400 ng / mL within two days of the first administration of the lipid binding protein-based complex.

[0845] 217. The method of any one of embodiments 213 to 216, wherein the dose is effective to reduce serum levels of ferritin by up to 700 ng / mL within two days of the first administration of the lipid-binding protein-based complex.

[0846] 218. The method of any one of embodiments 213 to 217, wherein the dose is effective to reduce serum levels of ferritin by at least 200 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0847] 219. The method of any one of embodiments 213 to 217, wherein the dose is effective to reduce serum levels of ferritin by at least 300 ng / mL within five days of the first administration of the lipid binding protein-based complex.

[0848] 220. The method of any one of embodiments 213 to 217, wherein the dose is effective to reduce serum levels of ferritin by at least 400 ng / mL within five days of the first administration of the lipid-binding protein-based complex.

[0849] 221. The method of any one of embodiments 213 to 220, wherein the dose is effective to reduce serum levels of ferritin by up to 700 ng / mL within two days of the first administration of the lipid-binding protein-based complex.

[0850] 222. The method of any one of embodiments 201 to 221, wherein the dose is effective to reduce serum levels of interleukin 8 (IL-8).

[0851] 223. The method of embodiment 222, wherein the dose is effective to reduce serum levels of IL-8 by at least 100 pg / mL within two days of the first administration of the lipid binding protein-based complex.

[0852] 224. The method of embodiment 222, wherein the dose is effective to reduce serum levels of IL-8 by at least 150 pg / mL within two days of the first administration of the lipid binding protein-based complex.

[0853] 225. The method of any one of embodiments 222 to 224, wherein the dose is effective to reduce serum levels of IL-8 by up to 300 pg / mL within two days of the first administration of the lipid binding protein-based complex.

[0854] 226. The method of any one of embodiments 222 to 225, wherein the dose is effective to reduce serum levels of IL-8 by at least 100 pg / mL within five days of the first administration of the lipid binding protein-based complex.

[0855] 227. The method of any one of embodiments 222 to 225, wherein the dose is effective to reduce serum levels of IL-8 by at least 150 pg / mL within five days of the first administration of the lipid binding protein-based complex.

[0856] 228. The method of any one of embodiments 222 to 227, wherein the dose is effective to reduce serum levels of IL-8 by up to 300 pg / mL within five days of the first administration of the lipid binding protein-based complex.

[0857] 229. The method of any one of embodiments 201 to 228, wherein the dose is effective to reduce serum levels of granulocyte-macrophage colony stimulating factor (GM-CSF).

[0858] 230. The method of any one of embodiments 201 to 229, wherein the dose is effective to reduce serum levels of monocyte chemoattractant protein (MCP) 1.

[0859] 231. The method of any one of embodiments 201 to 230, wherein the dose is effective to reduce serum levels of tumor necrosis factor alpha (TNF-α).

[0860] 232. The method of any one of embodiments 201 to 231, wherein the dose is effective to reduce serum levels of KIM-1.

[0861] 233. The method of any one of embodiments 201 to 232, wherein the dose is effective to reduce the serum level of the one or more inflammatory markers from an elevated range to a normal range.

[0862] 234. The method of any one of embodiments 201 to 233, wherein the dose is effective to reduce the serum level of the one or more inflammatory markers by at least 20%, at least 40%, or at least 60%.

[0863] 235. The method of any one of embodiments 1 to 234, wherein the subject has an endotoxin activity level of >0.6 prior to administration of the lipid binding protein-based complex.

[0864] 236. The method of any one of embodiments 1 to 235, wherein the dose is effective to reduce the level of endotoxin activity in the subject.

[0865] 237. The method of any one of embodiments 1 to 236, wherein the dose is effective to transiently increase serum triglyceride levels.

[0866] 238. A method as described in embodiment 237, wherein the transient increase is an increase of up to 9 days.

[0867] 239. The method of any one of embodiments 1 to 238, wherein the lipid binding protein-based complex is a reconstituted HDL or a HDL mimetic.

[0868] 240. The method of any one of embodiments 1 to 238, wherein the lipid binding protein-based complex is an apomer or a cargomer.

[0869] 241. The method of any one of embodiments 1 to 240, wherein the lipid binding protein-based complex comprises sphingomyelin.

[0870] 242. The method of any one of embodiments 1 to 241, wherein the lipid-binding protein-based complex comprises a negatively charged lipid.

[0871] 243. A method as described in embodiment 242, wherein the negatively charged lipid is 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol) (DPPG) or a salt thereof.

[0872] 244. A method as described in embodiment 239, wherein the lipid binding protein-based complex is CER-001, CSL-111, CSL-112, CER-522 or ETC-216.

[0873] 245. A method as described in embodiment 244, wherein the lipid binding protein-based complex is CER-001.

[0874] 246. The method of any one of embodiments 1 to 245, wherein the lipid binding protein-based complex is administered systemically, optionally by infusion.

[0875] 247. The method of any one of embodiments 1 to 246, wherein the lipid binding protein-based complex is administered until serum levels of one or more inflammatory markers decrease.

[0876] 248. The method of embodiment 247, wherein the lipid binding protein-based complex is administered until the serum level of one or more inflammatory markers decreases to a normal range.

[0877] 249. A method as described in embodiment 247, wherein the lipid-binding protein-based complex is administered until the serum level of one or more inflammatory markers decreases below the baseline level of the one or more inflammatory markers measured before the administration of the lipid-binding protein-based complex.

[0878] 250. The method of any one of embodiments 1 to 249, wherein each individual dose of the lipid-binding protein-based complex administered is 4-40 mg / kg (based on protein weight).

[0879] 251. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 4-30 mg / kg (based on protein weight).

[0880] 252. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 15-25 mg / kg (based on protein weight).

[0881] 253. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 10-30 mg / kg (based on protein weight).

[0882] 254. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 10-20 mg / kg (based on protein weight).

[0883] 255. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 5 mg / kg (based on protein weight).

[0884] 256. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 10 mg / kg (based on protein weight).

[0885] 257. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 15 mg / kg (based on protein weight).

[0886] 258. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 20 mg / kg (based on protein weight).

[0887] 259. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 5 to 15 mg / kg (based on protein weight).

[0888] 260. A method as described in embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 10 to 20 mg / kg (based on protein weight).

[0889] 261. The method of embodiment 250, wherein each individual dose of the lipid-binding protein-based complex is 15 to 25 mg / kg (based on protein weight).

[0890] 262. The method of any one of embodiments 1 to 261, wherein the dose is administered according to an induction regimen, optionally followed by a consolidation regimen.

[0891] 263. A method as described in embodiment 262, wherein the induction regimen comprises administering the lipid binding protein-based complex once a day or twice a day.

[0892] 264. The method of embodiment 262 or embodiment 263, wherein the consolidation regimen comprises administering the lipid binding protein-based complex once a day or once every two days.

[0893] 265. The method of any one of embodiments 1 to 264, wherein the subject is not treated with a maintenance regimen.

[0894] 266. The method of any one of embodiments 262 to 265, wherein the consolidation regimen comprises administering to the subject one or more doses of the lipid-binding protein-based complex one or more days after administration of the final dose of the induction regimen.

[0895] 267. A method as described in embodiment 266, wherein the first dose of the lipid binding protein-based complex administered during the consolidation regimen is administered two or more days after the final dose of the induction regimen.

[0896] 268. A method as described in embodiment 266, wherein the first dose of the lipid binding protein-based complex administered during the consolidation regimen is administered three or more days after the final dose of the induction regimen.

[0897] 269. A method as described in embodiment 268, wherein the first dose of the lipid binding protein-based complex administered during the consolidation regimen is administered three days after the final dose of the induction regimen.

[0898] 270. The method of any one of embodiments 262 to 269, comprising an induction regimen comprising administering the lipid-binding protein-based complex twice a day on days 1, 2, and 3, and a consolidation regimen comprising two doses of the lipid-binding protein-based complex on day 6.

[0899] 271. The method of any one of embodiments 262 to 270, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 4-40 mg / kg (based on protein weight).

[0900] 272. The method of any one of embodiments 262 to 271, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 4-30 mg / kg (based on protein weight).

[0901] 273. The method of any one of embodiments 262 to 271, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 15-25 mg / kg (based on protein weight).

[0902] 274. The method of any one of embodiments 262 to 271, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 10-30 mg / kg (based on protein weight).

[0903] 275. The method of any one of embodiments 262 to 271, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 10-20 mg / kg (based on protein weight).

[0904] 276. The method of any one of embodiments 262 to 271, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 5 mg / kg (based on protein weight).

[0905] 277. The method of any one of embodiments 262 to 271, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 10 mg / kg (based on protein weight).

[0906] 278. The method of any one of embodiments 262 to 271, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 15 mg / kg (based on protein weight).

[0907] 279. The method of any one of embodiments 262 to 271, wherein each individual dose of the lipid-binding protein-based complex administered in the induction regimen is 20 mg / kg (based on protein weight).

[0908] 280. The method of any one of embodiments 262 to 279, wherein the dose of the lipid-binding protein-based complex administered in the consolidation regimen is 5 mg / kg to 15 mg / kg (based on protein weight).

[0909] 281. The method of any one of embodiments 262 to 279, wherein the dose of the lipid-binding protein-based complex administered in the consolidation regimen is 10 mg / kg to 20 mg / kg (based on protein weight).

[0910] 282. The method of any one of embodiments 262 to 279, wherein the dose of the lipid-binding protein-based complex administered in the consolidation regimen is 15 mg / kg to 25 mg / kg (based on protein weight).

[0911] 283. The method of any one of embodiments 262 to 279, wherein the dose of the lipid-binding protein-based complex administered in the consolidation regimen is 5 mg / kg (based on protein weight).

[0912] 284. The method of any one of embodiments 262 to 279, wherein the dose of the lipid-binding protein-based complex administered in the consolidation regimen is 10 mg / kg (based on protein weight).

[0913] 285. The method of any one of embodiments 262 to 279, wherein the dose of the lipid-binding protein-based complex administered in the consolidation regimen is 15 mg / kg (based on protein weight).

[0914] 286. The method of any one of embodiments 1 to 285, wherein each individual dose of the lipid-binding protein-based complex administered is 300 mg to 4000 mg (based on protein weight).

[0915] 287. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 300 mg to 3000 mg (based on protein weight).

[0916] 288. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 300 mg to 1500 mg (based on protein weight).

[0917] 289. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 400 mg to 4000 mg (based on protein weight).

[0918] 290. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 400 mg to 1500 mg (based on protein weight).

[0919] 291. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 500 mg to 1200 mg (based on protein weight).

[0920] 292. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 500 mg to 1000 mg (based on protein weight).

[0921] 293. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 600 mg to 3000 mg (based on protein weight).

[0922] 294. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 800 mg to 3000 mg (based on protein weight).

[0923] 295. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 1000 mg to 2400 mg (based on protein weight).

[0924] 296. The method of embodiment 286, wherein each individual dose of the lipid-binding protein-based complex administered is 1000 mg to 2000 mg (based on protein weight).

[0925] 297. The method of any one of embodiments 1 to 296, wherein the high dose of the lipid-binding protein-based complex is 600 mg to 40 g (based on protein weight).

[0926] 298. A method as described in any one of embodiments 1 to 296, wherein the high dose of the lipid-binding protein-based complex is 3g to 35g (based on protein weight).

[0927] 299. The method of any one of embodiments 1 to 296, wherein the high dose of the lipid-binding protein-based complex is 5 g to 30 g (based on protein weight).

[0928] 300. The method of any one of embodiments 1 to 299, wherein the lipid binding protein-based complex is administered by infusion.

[0929] 301. The method of embodiment 300, wherein each individual dose is administered over a period of one to 24 hours.

[0930] 302. The method of embodiment 301, wherein each individual dose is administered within a 24 hour period.

[0931] 303. The method of any one of embodiments 1 to 302, further comprising administering an antihistamine to the subject prior to each individual dose.

[0932] 304. The method of embodiment 303, wherein the antihistamine comprises dexchlorpheniramine or hydroxyzine.

[0933] 305. The method of any one of embodiments 1 to 304, wherein the subject is receiving or has received one or more additional therapies and / or the method further comprises administering one or more additional therapies to the subject.

[0934] 306. The method of embodiment 305, wherein the one or more additional therapies comprise standard of care therapy.

[0935] 307. The method of embodiment 305 or embodiment 306, wherein the subject has an infection and the one or more additional therapies include antibiotic therapy.

[0936] 308. The method of embodiment 305 or embodiment 306, wherein the one or more additional therapies comprise hemodynamic support.

[0937] 309. The method of any one of embodiments 305 to 308, wherein the one or more additional therapies comprise one or more anti-IL-6 agents.

[0938] 310. The method of embodiment 309, wherein the one or more anti-IL-6 agents comprise tocilizumab, siltuximab, olotuzumab, ecitumomab, BMS-945429, sirukumab, levitra, CPSI-2364, or a combination thereof.

[0939] 311. The method of embodiment 310, wherein the one or more anti-IL-6 agents include tocilizumab.

[0940] 312. The method of any one of embodiments 305 to 311, wherein the one or more additional therapies comprise one or more corticosteroids.

[0941] 313. The method of embodiment 312, wherein the one or more corticosteroids comprise methylprednisolone, dexamethasone, or a combination thereof.

[0942] 314. The method of any one of embodiments 305 to 313, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise antibodies from a recovered COVID-19 patient.

[0943] 315. The method of any one of embodiments 305 to 314, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise antibodies to the spike protein of COVID-19.

[0944] 316. The method of any one of embodiments 305 to 315, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise one or more antiviral agents.

[0945] 317. The method of embodiment 316, wherein the one or more antiviral agents include lopinavir.

[0946] 318. The method of embodiment 316 or embodiment 317, wherein the one or more antiviral agents include remdesivir.

[0947] 319. The method of any one of embodiments 316 to 318, wherein the one or more antiviral agents comprises danoprevir.

[0948] 320. The method of any one of embodiments 316 to 319, wherein the one or more antiviral agents comprises galisvir.

[0949] 321. The method of any one of embodiments 316 to 320, wherein the one or more antiviral agents comprises darunavir.

[0950] 322. The method of any one of embodiments 316 to 321, wherein the one or more antiviral agents comprises ritonavir.

[0951] 323. The method of any one of embodiments 305 to 322, wherein the subject has or has had COVID-19 infection and the one or more additional therapies comprise chloroquine or hydroxychloroquine.

[0952] 324. The method of any one of embodiments 305 to 323, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprises azithromycin.

[0953] 325. The method of any one of embodiments 305 to 324, wherein the subject has or has had COVID-19 infection and the one or more additional therapies comprises interferon.

[0954] 326. The method of embodiment 325, wherein the interferon is interferon alpha.

[0955] 327. The method of embodiment 325, wherein the interferon is interferon beta.

[0956] 328. The method of any one of embodiments 325 to 327, wherein the interferon is pegylated.

[0957] 329. The method of any one of embodiments 1 to 328, wherein the lipid binding protein-based complex is CER-001.

[0958] 330. A method as described in embodiment 329, wherein CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids and the phospholipids sphingomyelin and DPPG, wherein the ApoA-I weight:total phospholipids weight ratio is 1:2.7+ / -20%, and wherein the sphingomyelin:DPPG weight:weight ratio is 97:3+ / -20%.

[0959] 331. A method as described in embodiment 329, wherein CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids and the phospholipids sphingomyelin and DPPG, wherein the ApoA-I weight:total phospholipids weight ratio is 1:2.7+ / -10%, and wherein the sphingomyelin:DPPG weight:weight ratio is 97:3+ / -10%.

[0960] 332. A method as described in embodiment 329, wherein CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids and the phospholipids sphingomyelin and DPPG, wherein the weight:weight ratio of ApoA-I:total phospholipids is 1:2.7, and wherein the weight:weight ratio of sphingomyelin:DPPG is 97:3.

[0961] 333. The method of any one of embodiments 330 to 332, wherein the ApoA-I has an amino acid sequence of amino acids 25-267 of SEQ ID NO:2.

[0962] 334. The method of any one of embodiments 330 to 333, wherein the ApoA-I is recombinantly expressed.

[0963] 335. The method of any one of embodiments 330 to 334, wherein CER-001 comprises a native sphingomyelin.

[0964] 336. A method as described in embodiment 335, wherein the natural sphingomyelin is egg sphingomyelin.

[0965] 337. The method of any one of embodiments 330 to 334, wherein CER-001 comprises a synthetic sphingomyelin.

[0966] 338. The method of embodiment 337, wherein the synthetic sphingomyelin is palmitoyl sphingomyelin.

[0967] 339. The method of any one of embodiments 329 to 338, wherein CER-001 is administered in the form of a formulation, wherein CER-001 is at least 95% homogeneous.

[0968] 340. The method of embodiment 339, wherein CER-001 is administered in the form of a formulation, wherein CER-001 is at least 97% homogeneous.

[0969] 341. The method of embodiment 339, wherein CER-001 is administered in the form of a formulation, wherein CER-001 is at least 98% homogeneous.

[0970] 342. The method of embodiment 339, wherein CER-001 is administered in the form of a formulation, wherein CER-001 is at least 99% homogeneous.

[0971] Various aspects of the present disclosure are described in the embodiments set forth in the following numbered paragraphs of Group 2.

[0972] Group 2:

[0973] 1. A method for treating a subject suffering from one or more symptoms associated with leukocytosis, the method comprising administering to the subject an ApoA-I formulation comprising apolipoprotein AI ("ApoA-I") and one or more lipids in an amount effective to reduce the subject's white blood cell count and / or improve one or more of the one or more symptoms associated with leukocytosis, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

[0974] 2. A method of treating a subject suffering from endothelial dysfunction, the method comprising administering to the subject an 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.

[0975] 3. A method of treating a subject having or at risk of developing carditis, the method comprising administering to the subject an 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.

[0976] 4. A method of treating a subject having or at risk of developing leukocytosis, the method comprising administering to the subject an ApoA-I preparation comprising ApoA-I and one or more lipids in an amount effective to reduce the subject's white blood cell count, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

[0977] 5. A method of treating a subject who is experiencing or has experienced an acute coronary syndrome or stroke, the method comprising administering to the subject an 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.

[0978] 6. The method of any one of embodiments 1 to 5, wherein the ApoA-I has an amino acid sequence of amino acids 25-267 of SEQ ID NO:2.

[0979] 7. The method of any one of embodiments 1 to 6, wherein the ApoA-I is recombinantly expressed.

[0980] 8. The method of any one of embodiments 1 to 7, wherein the one or more lipids comprise a neutral lipid.

[0981] 9. A method as described in embodiment 8, wherein the neutral lipid comprises sphingomyelin.

[0982] 10. The method of embodiment 9, wherein the neutral lipid consists of sphingomyelin.

[0983] 11. The method of embodiment 9 or embodiment 10, wherein the sphingomyelin comprises native sphingomyelin.

[0984] 12. The method of embodiment 11, wherein the natural sphingomyelin is hen egg sphingomyelin.

[0985] 13. The method of embodiment 9 or embodiment 10, wherein the sphingomyelin comprises a synthetic sphingomyelin.

[0986] 14. The method of embodiment 13, wherein the synthetic sphingomyelin is palmitoyl sphingomyelin.

[0987] 15. The method of any one of embodiments 1 to 10, wherein the one or more lipids further comprise a negatively charged lipid.

[0988] 16. The method of embodiment 15, wherein the negatively charged lipid comprises 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] ("DPPG") or a salt thereof.

[0989] 17. A method as described in embodiment 16, wherein the negatively charged lipid consists of DPPG or a salt thereof.

[0990] 18. The method of any one of embodiments 11 to 17, wherein the molar ratio of the negatively charged lipid to the neutral lipid to the component of ApoA-I in the formulation is 2-6:90-120:1.

[0991] 19. The method of any one of embodiments 8 to 18, wherein the lipids consist of 95% to 99% by weight neutral phospholipids and 1% to 5% by weight negatively charged phospholipids.

[0992] 20. The method of embodiment 19, wherein the lipids consist of 96 to 98 wt% neutral phospholipids and 2 to 4 wt% negatively charged phospholipids.

[0993] 21. The method of embodiment 20, wherein the lipids consist of 97 wt% neutral phospholipids and 3 wt% negatively charged phospholipids.

[0994] 22. The method of any one of embodiments 1 to 21, having a ratio of ApoA-I to lipid ranging from 1 :2 to 1 :3 by weight.

[0995] 23. The method of embodiment 22, having a ratio of ApoA-I to lipid of about 1:2.7 by weight.

[0996] 24. The method of any one of embodiments 1 to 23, wherein the lipoprotein complex is at least 95% homogeneous as reflected by a single peak in gel permeation chromatography.

[0997] 25. An 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, for use in treating a subject suffering from one or more symptoms associated with leukocytosis.

[0998] 26. An 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, for use in treating a subject suffering from endothelial dysfunction.

[0999] 27. An 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, for use in treating a subject having or at risk of developing carditis.

[1000] 28. An 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, for use in treating a subject having or at risk of developing leukocytosis.

[1001] 29. An 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, for use in treating a subject who is experiencing or has experienced an acute coronary syndrome or stroke.

[1002] 30. The formulation for use according to any one of embodiments 25 to 29, wherein the ApoA-I has an amino acid sequence of amino acids 25-267 of SEQ ID NO:2.

[1003] 31. The formulation for use according to any one of embodiments 25 to 30, wherein the ApoA-I is recombinantly expressed.

[1004] 32. The formulation for use according to any one of embodiments 25 to 31, wherein the one or more lipids comprise neutral lipids.

[1005] 33. The formulation for use according to embodiment 32, wherein the neutral lipid comprises sphingomyelin.

[1006] 34. The formulation for use according to embodiment 33, wherein the neutral lipids consist of sphingomyelin.

[1007] 35. The formulation for use according to embodiment 33 or embodiment 34, wherein the sphingomyelin comprises native sphingomyelin.

[1008] 36. The formulation for use according to embodiment 35, wherein the natural sphingomyelin is hen egg sphingomyelin.

[1009] 37. The formulation for use according to embodiment 33 or embodiment 34, wherein the sphingomyelin comprises a synthetic sphingomyelin.

[1010] 38. The formulation for use according to embodiment 37, wherein the synthetic sphingomyelin is palmitoyl sphingomyelin.

[1011] 39. The formulation for use according to any one of embodiments 25 to 34, wherein the one or more lipids further comprise negatively charged lipids.

[1012] 40. The formulation for use according to embodiment 39, wherein the negatively charged lipid comprises 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] ("DPPG") or a salt thereof.

[1013] 41. The formulation for use according to embodiment 40, wherein the negatively charged lipid consists of DPPG or a salt thereof.

[1014] 42. The formulation for use according to any one of embodiments 35 to 41, wherein the molar ratio of the negatively charged lipid to the neutral lipid to the component of ApoA-I in the formulation is 2-6:90-120:1.

[1015] 43. The formulation for use according to any one of embodiments 32 to 42, wherein the lipids consist of 95% to 99% by weight of neutral phospholipids and 1% to 5% by weight of negatively charged phospholipids.

[1016] 44. The method of embodiment 43, wherein the lipids consist of 96 to 98 wt% neutral phospholipids and 2 to 4 wt% negatively charged phospholipids.

[1017] 45. The formulation for use according to embodiment 44, wherein the lipids consist of 97% by weight of neutral phospholipids and 3% by weight of negatively charged phospholipids.

[1018] 46. ​​A formulation for use according to any one of embodiments 25 to 45, having a ratio of ApoA-I to lipid in the range of 1 :2 to 1 :3 by weight.

[1019] 47. The formulation for use according to embodiment 46, which has a ratio of ApoA-I to lipid of about 1:2.7 by weight.

[1020] 48. The formulation for use according to any one of embodiments 25 to 47, wherein the lipoprotein complex is at least 95% homogeneous as reflected by a single peak in gel permeation chromatography.

[1021] 49. An 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, for use in the preparation of a medicament for one or more symptoms associated with leukocytosis.

[1022] 50. An 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, for use in the preparation of a medicament for endothelial dysfunction.

[1023] 51. An 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, for use in the preparation of a medicament for use in carditis or at risk of developing carditis.

[1024] 52. An 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, for use in the preparation of a medicament for use in leukocytosis or the risk of developing leukocytosis.

[1025] 53. An 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, for use in the preparation of a medicament for acute coronary syndrome or stroke.

[1026] 54. The formulation for use according to any one of embodiments 49 to 53, wherein the ApoA-I has an amino acid sequence of amino acids 25-267 of SEQ ID NO:2.

[1027] 55. The formulation for use according to any one of embodiments 49 to 54, wherein the ApoA-I is recombinantly expressed.

[1028] 56. The formulation for use according to any one of embodiments 49 to 55, wherein the one or more lipids comprise neutral lipids.

[1029] 57. The formulation for use according to embodiment 56, wherein the neutral lipid comprises sphingomyelin.

[1030] 58. The formulation for use according to embodiment 57, wherein the neutral lipids consist of sphingomyelin.

[1031] 59. The formulation for use according to embodiment 57 or embodiment 58, wherein the sphingomyelin comprises native sphingomyelin.

[1032] 60. The formulation for use according to embodiment 59, wherein the natural sphingomyelin is hen egg sphingomyelin.

[1033] 61. The formulation for use according to embodiment 57 or embodiment 58, wherein the sphingomyelin comprises a synthetic sphingomyelin.

[1034] 62. The formulation for use according to embodiment 61, wherein the synthetic sphingomyelin is palmitoyl sphingomyelin.

[1035] 63. The formulation for use according to any one of embodiments 49 to 58, wherein the one or more lipids further comprise negatively charged lipids.

[1036] 64. The formulation for use according to embodiment 63, wherein the negatively charged lipid comprises 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] ("DPPG") or a salt thereof.

[1037] 65. The formulation for use according to embodiment 64, wherein the negatively charged lipid consists of DPPG or a salt thereof.

[1038] 66. The formulation for use according to any one of embodiments 59 to 65, wherein the molar ratio of the negatively charged lipid to the neutral lipid to the component of ApoA-I in the formulation is 2-6:90-120:1.

[1039] 67. The formulation for use according to any one of embodiments 56 to 66, wherein the lipids consist of 95% to 99% by weight of neutral phospholipids and 1% to 5% by weight of negatively charged phospholipids.

[1040] 68. A method as described in embodiment 67, wherein the lipids are composed of 96% to 98% by weight neutral phospholipids and 2% to 4% by weight negatively charged phospholipids.

[1041] 69. The formulation for use according to embodiment 68, wherein the lipids consist of 97% by weight of neutral phospholipids and 3% by weight of negatively charged phospholipids.

[1042] 70. The formulation for use according to any one of embodiments 49 to 69, having a ratio of ApoA-I to lipid in the range of 1 :2 to 1 :3 by weight.

[1043] 71. The formulation for use according to embodiment 70, which has a ratio of ApoA-I to lipid of about 1:2.7 by weight.

[1044] 72. The formulation for use according to any one of embodiments 49 to 71, wherein the lipoprotein complex is at least 95% homogeneous as reflected by a single peak in gel permeation chromatography.

[1045] 73. The method or formulation for use according to any one of embodiments 1 to 72, wherein the dose is administered twice a day.

[1046] 74. The method or formulation for use according to any one of embodiments 1 to 73, wherein the dose is administered for five days.

[1047] 75. The method or formulation for use according to any one of embodiments 1 to 74, wherein the dose is effective to reduce the white blood cell count of the subject by at least 2000 WBC / microliter within five days of the first administration of the lipid binding protein-based complex.

[1048] 76. The method or formulation for use according to any one of embodiments 1 to 75, wherein the dose is effective to reduce the white blood cell count of the subject by at least 3000 WBC / microliter within five days of the first administration of the lipid binding protein-based complex.

Claims

1. A method of treating a subject suffering from one or more symptoms associated with leukocytosis, the method comprising administering to the subject a lipid binding protein-based complex in an amount effective to reduce the subject's white blood cell count and / or improve one or more of the one or more symptoms associated with leukocytosis.

2. The method of claim 1, wherein the one or more symptoms include fever; bleeding or bruising; sweating; pain or tingling in the legs, arms, or abdomen; one or more vision problems; unclear thinking; loss of appetite; and / or difficulty breathing (e.g., shortness of breath, lower than normal blood oxygen levels).

3. The method of claim 1 or claim 2, wherein the dosage of the lipid binding protein-based complex is effective to improve one or more of the one or more symptoms associated with the leukocytosis.

4. The method of any one of claims 1 to 3, wherein the subject has or is at risk of developing leukocytosis due to inflammation, infection, a leukocyte disorder, physical stress, emotional stress, a drug, or an allergic reaction.

5. The method of any one of claims 1 to 4, wherein the subject has an infection.

6. The method of claim 5, wherein the infection is a viral infection, optionally wherein the viral infection is a coronavirus infection, the coronavirus optionally being COVID-19; a bacterial infection; a fungal infection; or a parasitic infection.

7. The method of any one of claims 1 to 6, wherein the subject suffers from diabetes.

8. The method of any one of claims 1 to 7, wherein the subject has a leukocyte disorder.

9. The method of any one of claims 1 to 8, wherein the subject suffers from leukocytosis.

10. The method of claim 9, wherein the subject has a serotype greater than 11 x 10 9 / L white blood cell count.

11. The method of any one of claims 1 to 10, wherein the subject is at risk for developing leukocytosis.

12. A method of treating a subject suffering from endothelial dysfunction, the method comprising administering to the subject a dose of a lipid binding protein-based complex, wherein the dose is a high dose.

13. The method of claim 12, wherein the dose is effective to improve vascular endothelial function in the subject, optionally wherein vascular endothelial function is measured by circulating VCAM-1 and / or ICAM-1.

14. The method of claim 13, wherein the dose is effective to reduce circulating VCAM-1 in the subject by at least 100 ng / mL within two days of the first administration of the lipid binding protein-based complex.

15. The method of claim 13 or claim 14, wherein the dose is effective to reduce circulating VCAM-1 in the subject by up to 400 ng / mL within five days of the first administration of the lipid binding protein-based complex.

16. The method of any one of claims 13 to 15, wherein the dose is effective to reduce circulating ICAM-1 in the subject by at least 50 ng / mL within two days of the first administration of the lipid binding protein-based complex.

17. The method of any one of claims 13 to 16, wherein the dose is effective to reduce circulating ICAM-1 in the subject by up to 125 ng / mL within five days of the first administration of the lipid binding protein-based complex.

18. A method of treating a subject having or at risk of developing carditis, the method comprising administering to the subject a dose of a lipid binding protein-based complex, optionally wherein the carditis is myocarditis and / or pericarditis.

19. The method of claim 18, wherein the subject has an infection, such as a viral infection, optionally a coronavirus infection, optionally COVID-19; or a bacterial infection.

20. The method of claim 18 or claim 19, wherein the dose of the lipid binding protein-based complex is effective to improve one or more symptoms of the carditis.

21. The method of any one of claims 18 to 20, wherein the subject is at risk for carditis.

22. The method of claim 21, wherein the dosage of the lipid binding protein-based complex is effective to prevent the carditis or reduce the severity of the carditis.

23. The method of any one of claims 1 to 22, wherein the dose is effective to reduce circulating VCAM-1 and / or ICAM-1 in the subject.

24. A method of treating a subject having or at risk of developing leukocytosis, the method comprising administering to the subject an amount of a lipid binding protein-based complex effective to reduce the subject's white blood cell count.

25. A method of treating a subject who is experiencing or has experienced an acute coronary syndrome or stroke, the method comprising administering to the subject a dose of a lipid binding protein-based complex, wherein the dose is a high dose.

26. The method of claim 25, wherein the subject is experiencing or has experienced an acute coronary syndrome, optionally wherein the acute coronary syndrome is myocardial infarction (e.g., ST-segment elevation myocardial infarction or non-ST-segment elevation myocardial infarction) or unstable angina.

27. The method of claim 25, wherein the subject is experiencing or has experienced a stroke.

28. A method of treating a subject suffering from one or more symptoms associated with leukocytosis, the method comprising administering to the subject an ApoA-I formulation comprising apolipoprotein AI ("ApoA-I") and one or more lipids in an amount effective to reduce the subject's white blood cell count and / or improve one or more of the one or more symptoms associated with leukocytosis, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

29. A method of treating a subject suffering from endothelial dysfunction, the method comprising administering to the subject an 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.

30. A method of treating a subject having or at risk of developing carditis, the method comprising administering to the subject an 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.

31. A method of treating a subject having or at risk of developing leukocytosis, the method comprising administering to the subject an ApoA-I formulation comprising ApoA-I and one or more lipids in an amount effective to reduce the subject's white blood cell count, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

32. A method of treating a subject who is experiencing or has experienced an acute coronary syndrome or stroke, the method comprising administering to the subject an 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.

33. The method of any one of claims 28 to 32, wherein the ApoA-I has an amino acid sequence of amino acids 25-267 of SEQ ID NO:

2.

34. The method of any one of claims 28 to 33, wherein the ApoA-I is recombinantly expressed.

35. The method of any one of claims 28 to 34, wherein the one or more lipids comprise a neutral lipid.

36. The method of claim 35, wherein the neutral lipid comprises sphingomyelin.

37. The method of claim 36, wherein the neutral lipid consists of sphingomyelin.

38. The method of claim 36 or claim 37, wherein the sphingomyelin comprises native sphingomyelin.

39. The method of claim 38, wherein the natural sphingomyelin is hen egg sphingomyelin.

40. The method of claim 36 or claim 37, wherein the sphingomyelin comprises a synthetic sphingomyelin.

41. The method of claim 40, wherein the synthetic sphingomyelin is palmitoyl sphingomyelin.

42. The method of any one of claims 28 to 37, wherein the one or more lipids further comprise a negatively charged lipid.

43. The method of claim 42, wherein the negatively charged lipid comprises 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] ("DPPG") or a salt thereof.

44. The method of claim 43, wherein the negatively charged lipid consists of DPPG or a salt thereof.

45. The method of any one of claims 38 to 44, wherein the molar ratio of the negatively charged lipid to the neutral lipid to the component of ApoA-I in the formulation is 2-6:90-120:

1.

46. ​​The method of any one of claims 35 to 45, wherein the lipids consist of 95 to 99% by weight of neutral phospholipids and 1 to 5% by weight of negatively charged phospholipids.

47. The method of claim 46, wherein the lipids consist of 96 to 98 weight percent neutral phospholipids and 2 to 4 weight percent negatively charged phospholipids.

48. The method of claim 47, wherein the lipids consist of 97% by weight neutral phospholipids and 3% by weight negatively charged phospholipids.

49. The method of any one of claims 28 to 48, having a ratio of ApoA-I to lipid ranging from 1 :2 to 1 :3 by weight.

50. The method of claim 49, having a ratio of ApoA-I to lipid of about 1 :2.7 by weight.

51. The method of any one of claims 28 to 50, wherein the lipoprotein complex is at least 95% homogeneous as reflected by a single peak in gel permeation chromatography.

52. The method of any one of claims 1 to 51, wherein the dose is administered twice daily.

53. The method of any one of claims 1 to 52, wherein the dose is administered for five days.

54. The method of any one of claims 1 to 53, wherein the dose is effective to reduce the subject's white blood cell count by at least 2000 WBC / microliter within five days of the first administration of the lipid binding protein-based complex.

55. The method of any one of claims 1 to 54, wherein the dose is effective to reduce the subject's white blood cell count by at least 3000 WBC / microliter within five days of the first administration of the lipid binding protein-based complex.

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