Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides
  • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
  • A61K38/1703—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
  • A61K38/1709—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
  • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/0012—Galenical forms characterised by the site of application
  • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/12—Antivirals
  • A61P31/14—Antivirals for RNA viruses
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P37/00—Drugs for immunological or allergic disorders
  • A61P37/02—Immunomodulators
  • A61P37/06—Immunosuppressants, e.g. drugs for graft rejection

Definitions

• Common signs and symptoms of sepsis include fever, increased heart rate, increased breathing rate, and confusion. There may also be symptoms related to a specific infection, such as a cough with pneumonia, or painful urination with a kidney infection (Jui et al., 2011, “Ch.146: Septic Shock.” In Tintinalli JE, et al. (eds.). Tintinalli's Emergency Medicine: A Comprehensive Study Guide (7th ed.). New York: McGraw-Hill. pp.1003– 14). Severe sepsis can be associated with poor organ function or blood flow (Dellinger et al., 2013, Critical Care Medicine.41(2):580–637).
• a sepsis diagnosis can be based on the shortened sequential organ failure assessment score (SOFA score), also known as the quick SOFA score (qSOFA), which requires at least two of the following three: increased breathing rate, change in the level of consciousness, and low blood pressure (Singer et al., 2016, JAMA.315(8):801–10).
• SOFA score also known as the quick SOFA score (qSOFA)
• qSOFA quick SOFA score
• Sepsis can require immediate treatment with intravenous fluids and antimicrobials (Rhodes et al., 2017, Intensive Care Medicine.43(3):304–377). Ongoing care often continues in an intensive care unit. If an adequate trial of fluid replacement is not enough to maintain blood pressure, then the use of medications that raise blood pressure can become necessary.
• Acute kidney injury is a common occurrence in ICU patients, with an estimated incidence of >50% (Hoste et al., 2015, Intensive Care Med; 41:1411–1423). Furthermore, increasing AKI severity is associated with increased mortality. Sepsis is the major cause of AKI, accounting for 45% to 70% of cases, and approximately 25% of sepsis is of intra-abdominal origin (Seymour et al., 2016, JAMA, 315:762–774; Bagshaw et al., 2007, Clin J Am Soc Nephrol, 2:431– 439).
• IRI Ischemia/reperfusion injury
• CSA AKI Cardiac surgery associated AKI
• Post-surgical IL6 and IL10 levels are predictive of AKI development and outcome (Zhang et al., 2015, J Am Soc Nephrol.26(12):3123-32) and there are no good treatment options other than dialysis (Küllmar et al., 2020, Crit Care Clin.36(4):691-704. [0009] Early diagnosis of AKI in the setting of sepsis is important in order to provide optimal treatment and avoid further kidney injury (Peerapornratana et al., 2019, Kidney International 2019, 96:1083–1099). Treatment options for sepsis-related AKI are limited to supportive care.
• ATII showed some benefit in a post-hoc analysis of AKI patients in a high-output shock study (ATHOS-3) and is currently being study in sepsis-related AKI in the ASK-IT trial (NCT00711789), however no updates have been given since 2011.
• Levocarnitine did not show organ dysfunction improvement in septic shock in the RACE study (Jones et al., 2018, JAMA network open, 1:e186076) but is currently being studied as an adjunct treatment for septic shock patients with AKI in the Carnisave trial (NCT02664753).
• High- density lipoprotein (HDL) is a key component of circulating blood and mainly contains phospholipids, free cholesterol, cholesteryl ester, triglycerides, apolipoproteins (Apo A-I, Apo A- II), and other proteins.
• HDL vascular endothelial function and immunity
• HDL high levels have been associated with increased risk of acute kidney injury (AKI) in course of sepsis (Roveran et al., 2017, Journal of internal medicine, 281:518–529; Zhang et al., 2009, Am J Physiol Heart Circ Physiol 297:H866–H873). Renal function and plasma HDL are strongly related to each other as kidneys are implicated in the recycling of senescent HDL particles and their filtration function is associated with their levels and contents (Yang et al., 2016, Current opinion in nephrology and hypertension, 25:174–179).
• rHDL recombinant HDL
• CSL-111 a rHDL originally produced for atherosclerosis treatment (Tardif et al., 2007, JAMA, 297(15):1675-82), has shown efficacy in reducing the inflammatory response during LPS-induced endotoxemia in vitro and in rabbit (Casas et al.,1995, The Journal of surgical research, 59:544– 552) and human models (Pajkrt et al., 1996, Journal of Experimental Medicine, 184(5):1601-8; Pajkrt et al., 1997, Thrombosis and Haemostasis, 77(2):303-7).
• HDL and HDL mimetic include human immunodeficiency virus (HIV) and hepatitis C virus (HCV).
• HCV human immunodeficiency virus
• HCV hepatitis C virus
• no HDL or HDL mimetic has received regulatory approval for the treatment of sepsis or AKI, including sepsis-related AKI, ischemia/reperfusion AKI and CSA AKI. 3.3.
• Cytokine release syndrome also called cytokine storm syndrome (CSS) is a systemic inflammatory response that can be caused by a variety of factors such as infection or treatment with some types of immunotherapy, such as monoclonal antibodies and adoptive T- cell therapies (Shimabukuro-Vornhagen, et al., 2018, J. Immunotherapy Cancer, 6:56). Symptoms of CRS include fever, nausea, headache, rash, rapid heartbeat, low blood pressure, and trouble breathing.
• CRS interleukin-6
• CRS may contribute to death of these patients (Zhang et al., 2020, International Journal of Antimicrobial Agents, doi.org/10.1016/j.ijantimicag.2020.105954; Mehta et al., 2020, The Lancet, 395(10229):1033- 1034).
• AKI including sepsis related AKI, ischemia/reperfusion AKI and CSA AKI
• CRS for example CRS associated with immunotherapy and CRS secondary to infections such as COVID-19. 4.
• the present disclosure provides methods for treating subjects with acute conditions, for example conditions associated with acute inflammation, with a high dose of a lipid binding protein-based complex.
• the high dose is typically higher than a dose that would be used to treat a chronic condition, such as familial hypercholesterolemia.
• the high dose is typically administered over a relatively short period of time, for example, over a period of three days to two weeks, and typically comprises multiple administrations of the lipid binding protein-based complex, for example three to 10 individual doses.
• the individual doses can be separated by less than one day (e.g., twice daily administration), or one day or more (e.g., once daily administration).
• the lipid binding protein-based complex comprises a sphingomyelin and/or a negatively charged lipid, for example CER-001.
• CER-001 is a negatively charged lipoprotein complex, and comprises recombinant human ApoA-I, sphingomyelin (SM), and 1, 2-dihexadecanoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (Dipalmitoylphosphatidyl-glycerol; DPPG). It mimics natural, nascent discoidal pre-beta HDL, which is the form that HDL particles take prior to acquiring cholesterol.
• CER-001 therapy can reduce serum levels of inflammatory cytokines such as IL-6, thereby providing a clinical benefit to subjects having an acute condition or at risk of an acute condition, for example subjects having or at risk of an acute inflammatory condition.
• the present disclosure provides methods for treating subjects with sepsis and methods for treating subjects with AKI or at risk for AKI with lipid binding protein- based complexes (e.g., CER-001).
• the disclosure provides a method of treating a subject with sepsis, comprising administering to the subject a lipid binding protein-based complex (e.g., CER-001).
• the disclosure provides a method of treating a subject with acute kidney injury (AKI) or at risk for AKI (e.g., a subject with sepsis which has not yet caused AKI, an organ transplant recipient, or a subject who has undergone cardiac surgery, or a subject having acute or chronic liver disease and at risk of hepatorenal syndrome (HRS)), comprising administering to the subject a lipid binding protein-based complex (e.g., CER-001).
• a lipid binding protein-based complex e.g., CER-001
• the disclosure provides methods for treating cytokine release syndrome (CRS) and/or reducing one or more inflammatory markers in a subject in need thereof with a lipid binding protein-based complex (e.g., CER-001).
• the disclosure provides methods of treating a subject with CRS or at risk of CRS, e.g., a subject with CRS secondary to COVID-19 or a subject with CRS caused by immunotherapy, comprising administering a therapeutically effective amount of a lipid binding protein-based complex (e.g., CER-001) to the subject.
• a lipid binding protein-based complex e.g., CER-001
• the disclosure provides methods of reducing serum levels of one or more inflammatory markers, e.g., one or more markers associated with CRS such as IL-6, in a subject in need thereof.
• the subject can be, for example, a subject with CRS or a subject at risk of CRS, for example a subject infected with a virus such as COVID-19 or a subject receiving immunotherapy.
• the present disclosure provides dosing regimens for lipid binding protein-based therapy (e.g., CER-001 therapy) for subjects with an acute condition (e.g., associated with acute inflammation), for example sepsis, AKI (e.g., AKI caused by sepsis, ischemia/reperfusion, cardiac surgery, or hepatorenal syndrome), or at risk for an acute condition such as AKI (e.g., a subject with sepsis which has not yet led to AKI) or CRS.
• an acute condition e.g., associated with acute inflammation
• AKI e.g., AKI caused by sepsis, ischemia/reperfusion, cardiac surgery, or hepatorenal syndrome
• AKI e.g., a subject with sepsis which has not
• the dosing regimens of the disclosure typically entail multiple administrations of CER- 001 to a subject (e.g., administered daily).
• the CER-001 therapy can be continued for a pre- determined period, e.g., for one week or a period longer than one week (e.g., two weeks).
• administration of CER-001 to a subject can be continued until one or more symptoms of the acute condition (e.g., acute inflammation or CRS) are reduced or continued until the serum levels of one or more inflammatory markers are reduced, for example reduced to a normal level or reduced relative to a baseline measurement taken prior to the start of CER- 001 therapy.
• the acute condition e.g., acute inflammation or CRS
• the therapy can in some embodiments be continued until the subject has recovered from the infection or discontinues immunotherapy.
• the dosing regimens of the disclosure can entail administering a lipid binding protein- based complex (e.g., CER-001) to a 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.
• the induction regimen typically comprises administering multiple doses of the lipid binding protein-based complex (e.g., CER-001) to the subject, for example six doses over three days.
• the consolidation regimen typically comprises administering one or more doses of a lipid binding protein-based complex (e.g., CER-001) to the subject following the final dose of the induction regimen, for example one or more days after the final dose of the induction regimen.
• the first dose of the consolidation regimen is administered on the third day after the final dose of the induction regimen.
• a dosing regimen can comprise administration of a lipid binding protein-based complex (e.g., CER-001) to a subject according to an induction regimen on days 1, 2, and 3, and administration of the lipid binding protein-based complex to the subject according to a consolidation regimen on day 6.
• the consolidation regimen comprises two doses of the lipid binding protein-based complex.
• the disclosure provides methods of treating a subject having CRS, sepsis or AKI, or a subject at risk of CRS or AKI (e.g., a subject with COVID-19) with a lipid binding protein-based complex (e.g., CER-001) according to a dosage regimen comprising: - 2 doses per day on days 1, 2, and 3 (induction regimen) optionally followed by - 2 subsequent doses on day 4 or later (consolidation regimen).
• the regimen comprises: - 2 doses per day on days 1, 2, and 3 (induction regimen) followed by - 2 doses on day 6 (consolidation regimen).
• a lipid binding protein-based complex (e.g., CER-001) is administered in combination with a standard of care therapy for sepsis such as antibiotic therapy and/or hemodynamic support.
• a standard of care therapy for sepsis such as antibiotic therapy and/or hemodynamic support.
• an antihistamine e.g., dexchlorpheniramine, hydroxyzine, diphenhydramine, cetirizine, fexofenadine, or loratadine
• the antihistamine can reduce the likelihood of allergic reactions. 5.
• FIG.1 shows IL-6 serum levels in a pig model of sepsis-induced AKI (Example 1).
• FIG.2 shows soluble VCAM-1 serum levels in a pig model of sepsis-induced AKI (Example 2).
• FIG.3 shows soluble ICAM-1 serum levels in a pig model of sepsis-induced AKI (Example 3).
• FIG.4 shows LPS serum levels in a pig model of sepsis-induced AKI (Example 1).
• FIG.5 shows a schematic of the clinical study of Example 2.
• FIG.6 is a flowsheet for the study of Example 3.
• FIG.7 is a flowsheet for the study of Example 4. 6.
• the present disclosure provides methods for treating subjects with acute conditions, for example, an acute condition comprising acute inflammation, with a high dose of a lipid binding protein-based complex.
• the disclosure provides methods for treating subjects having sepsis using a lipid binding protein-based complex (e.g., CER-001).
• the disclosure provides methods for treating subjects with acute kidney injury (AKI) or at risk of AKI (e.g., due to sepsis, viral infection, ischemia/reperfusion, cardiac surgery, or hepatorenal syndrome) using a lipid binding protein-based complex (e.g., CER- 001).
• AKI acute kidney injury
• CER- 001 lipid binding protein-based complex
• the disclosure provides methods of treating a subject with CRS or at risk of CRS, e.g., a subject with CRS secondary to COVID-19 or a subject with CRS caused by immunotherapy.
• 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.
• Exemplary features of lipid binding protein-based complexes that can be used in the methods and compositions of the disclosure are described in Section 6.1. Exemplary subject populations who can be treated by the methods of the disclosure and with the compositions of the disclosure are described in Section 6.2.
• methods of the disclosure comprise administering a lipid binding protein-based complex (e.g., CER-001) to a subject in two phases.
• the 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.
• a lipid binding protein-based complex (e.g., CER-001) can be administered to a subject in a single phase, for example according to an administration regimen corresponding to the dose and administration frequency of an induction or consolidation regimen described herein.
• Induction regimens that can be used in the methods of the disclosure are described in Section 6.3 and consolidation regimens that can be used in the methods of the disclosure are described in Section 6.3.2.
• the dosing regimens of the disclosure comprise administering a lipid binding protein-based complex (e.g., CER-001) as monotherapy or as part of a combination therapy with one or more medications, for example in combination with a standard of care therapy for sepsis such as antibiotic treatment and/or hemodynamic support.
• Combination therapies are described in Section 6.4.
• the lipid binding protein-based complexes comprise HDL or HDL mimetic- based complexes.
• complexes can comprise a lipoprotein complex as described in U.S.
• Lipoprotein complexes can 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 apolipoproteins, peptides, or apolipoprotein peptide analogs or mimetics, for example one or more lipid binding protein molecules described in Section 6.1.2.
• the lipid fraction typically includes one or more phospholipids which can be neutral, negatively charged, positively charged, or a combination thereof. Exemplary phospholipids and other amphipathic molecules which can be included in the lipid fraction are described in Section 6.1.3.
• the lipid fraction contains at least one neutral phospholipid (e.g., a sphingomyelin (SM)) and, optionally, one or more negatively charged phospholipids.
• SM sphingomyelin
• the neutral and negatively charged phospholipids can have fatty acid chains with the same or different number of carbons and the same or different degree of saturation.
• the neutral and negatively charged phospholipids will have the same acyl tail, for example a C16:0, or palmitoyl, acyl chain.
• the weight ratio of the apolipoprotein fraction: lipid fraction ranges from about 1:2.7 to about 1:3 (e.g., 1:2.7).
• Any phospholipid that bears at least a partial negative charge at physiological pH can be used as the negatively charged phospholipid.
• Non-limiting examples include negatively charged forms, e.g., salts, of phosphatidylinositol, a phosphatidylserine, a phosphatidylglycerol and a phosphatidic acid.
• the negatively charged phospholipid is 1,2- dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)], or DPPG, a phosphatidylglycerol.
• Preferred salts include potassium and sodium salts.
• a lipoprotein complex used in the methods of the disclosure is a lipoprotein complex as described in U.S. Patent No.8,206,750 or WO 2012/109162 (and its U.S. counterpart, US 2012/0232005), the contents of each of which are incorporated herein in its entirety by reference.
• the protein component of the lipoprotein complex is as described in Section 6.1 and preferably in Section 6.1.1 of WO 2012/109162 (and US 2012/0232005), the lipid component is as described in Section 6.2 of WO 2012/109162 (and US 2012/0232005), which can optionally be complexed together in the amounts described in Section 6.3 of WO 2012/109162 (and US 2012/0232005).
• Section 6.1 preferably in Section 6.1.1 of WO 2012/109162 (and US 2012/0232005)
• the lipid component is as described in Section 6.2 of WO 2012/109162 (and US 2012/0232005)
• the contents of each of these sections are incorporated by reference herein.
• a lipoprotein complex of the disclosure is 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 by reference herein.
• a lipoprotein complex that can be used in the methods of the disclosure comprises 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 50-80 molecules of lecithin and 20-50 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure comprises 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 50 molecules of lecithin and 50 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure comprises 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 80 molecules of lecithin and 20 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure comprises 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 70 molecules of lecithin and 30 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure comprises 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 60 molecules of lecithin and 40 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure consists essentially of 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 50-80 molecules of lecithin and 20-50 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure consists essentially of 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 50 molecules of lecithin and 50 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure consists essentially of 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 80 molecules of lecithin and 20 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure consists essentially of 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 70 molecules of lecithin and 30 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure consists essentially of 2-4 ApoA-I equivalents, 2 molecules of charged phospholipid, 60 molecules of lecithin and 40 molecules of SM.
• a lipoprotein complex that can be used in the methods of the disclosure consists essentially of about 90 to 99.8 wt % lecithin and about 0.2 to 10 wt % negatively charged phospholipid, for example, about 0.2-1 wt %, 0.2-2 wt %, 0.2-3 wt %, 0.2-4 wt %, 0.2-5 wt %, 0.2-6 wt %, 0.2-7 wt %, 0.2-8 wt %, 0.2-9 wt % or 0.2-10 wt % total negatively charged phospholipid(s).
• HDL-based or HDL mimetic-based complexes can include a single type of lipid-binding protein, or mixtures of two or more different lipid-binding proteins, which may be derived from the same or different species.
• the complexes will preferably comprise lipid-binding proteins that are derived from, or correspond in amino acid sequence to, the animal species being treated, in order to avoid inducing an immune response to the therapy.
• lipid-binding proteins of human origin are preferably used for treatment of human patients.
• the use of peptide mimetic apolipoproteins may also reduce or avoid an immune response.
• the lipid component includes two types of phospholipids: a sphingomyelin (SM) and a negatively charged phospholipid.
• SM sphingomyelin
• Exemplary SMs and negatively charged lipids are described in Section 6.1.3.1.
• Lipid components including SM can optionally include small quantities of additional lipids. Virtually any type of lipids may be used, including, but not limited to, lysophospholipids, galactocerebroside, gangliosides, cerebrosides, glycerides, triglycerides, and cholesterol and its derivatives.
• the optional lipids When included, such optional lipids will typically comprise less than about 15 wt% of the lipid fraction, although in some instances more optional lipids could be included. 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.
• the phospholipid fraction contains egg SM or palmitoyl SM or phytosphingomyelin and DPPG in 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.
• the molar ratio of the lipid component to the protein component of complexes of the disclosure can vary, and will depend upon, among other factors, the identity(ies) of the apolipoprotein comprising the protein component, the identities and quantities of the lipids comprising the lipid component, and the desired size of the complex. Because the biological activity of apolipoproteins such as ApoA-I are thought to be mediated by the amphipathic helices comprising the apolipoprotein, it is convenient to express the apolipoprotein fraction of the lipid:apolipoprotein molar ratio using ApoA-I protein equivalents.
• ApoA-I contains 6-10 amphipathic helices, depending upon the method used to calculate the helices.
• Other apolipoproteins can be expressed in terms of ApoA-I equivalents based upon the number of amphipathic helices they contain.
• ApoA-IM which typically exists as a disulfide-bridged dimer, can be expressed as 2 ApoA-I equivalents, because each molecule of ApoA-IM contains twice as many amphipathic helices as a molecule of ApoA-I.
• a peptide apolipoprotein that contains a single amphipathic helix can be expressed as a 1/10-1/6 ApoA-I equivalent, because each molecule contains 1/10-1/6 as many amphipathic helices as a molecule of ApoA-I.
• the lipid:ApoA-I equivalent molar ratio of the lipoprotein complexes (defined herein as “Ri”) will range from about 105:1 to 110:1.
• the Ri is about 108:1. Ratios in weight can be obtained using a MW of approximately 650-800 for phospholipids.
• the molar ratio of lipid : ApoA-I equivalents ranges from about 80:1 to about 110:1, e.g., about 80:1 to about 100:1.
• the RSM for complexes can be about 82:1.
• lipoprotein complexes used in the methods of the disclosure are negatively charged complexes which comprise a protein fraction which is preferably mature, full-length ApoA-I, and a lipid fraction comprising a neutral phospholipid, sphingomyelin (SM), and negatively charged phospholipid.
• SM sphingomyelin
• the lipid component contains SM (e.g., egg SM, palmitoyl SM, phytoSM, or a combination thereof) and negatively charged phospholipid (e.g., DPPG) in a weight ratio (SM : negatively charged phospholipid) ranging from 90:10 to 99:1, more preferably ranging from 95:5 to 98:2, e.g., 97:3.
• SM negatively charged phospholipid
• the ratio of the protein component to lipid component can range from about 1:2.7 to about 1:3, with 1:2.7 being preferred. This corresponds to molar ratios of ApoA-I protein to lipid ranging from approximately 1:90 to 1:140.
• 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.
• the complex comprises CER-001, CSL-111, CSL-112, CER- 522 or ETC-216.
• the complex is CER-001.
• CER-001 as used in the literature and in the Examples below refers to a complex described in Example 4 of WO 2012/109162.
• WO 2012/109162 refers to CER-001 as a complex having a 1:2.7 lipoprotein weight:total phospholipid weight ratio with a SM:DPPG weight:weight ratio of 97:3.
• Example 4 of WO 2012/109162 also describes a method of its manufacture.
• CER-001 refers to a lipoprotein complex whose individual constituents can vary from CER-001 as described in Example 4 of WO 2012/109162 by up to 20%.
• the constituents of the lipoprotein complex vary from CER-001 as described in Example 4 of WO 2012/109162 by up to 10%.
• the constituents 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 can be natural or synthetic.
• the SM is a natural SM, for example a natural SM described in WO 2012/109162, e.g., chicken egg SM.
• the SM is a synthetic SM, for example a synthetic SM described in WO 2012/109162, e.g., synthetic palmitoylsphingomyelin, for example as described in WO 2012/109162. Methods for synthesizing palmitoylsphingomyelin are known in the art, for example as described in WO 2014/140787.
• the lipoprotein in CER-001, apolipoprotein A-I preferably has an amino acid sequence corresponding to amino acids 25 to 267 of SEQ ID NO:1 of WO 2012/109162.
• ApoA- I can be purified by animal sources (and in particular from human sources) or produced recombinantly.
• the ApoA-I in CER-001 is recombinant ApoA-I.
• CER- 001 used in a dosing regimen of the disclosure is preferably highly homogeneous, for example 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.
• CSL-111 is a reconstituted human ApoA-I purified from plasma complexed with soybean phosphatidylcholine (SBPC) (Tardif et al., 2007, JAMA 297:1675-1682).
• CSL-112 is a formulation of ApoA-I purified from plasma and reconstituted to form HDL suitable for intravenous infusion (Diditchenko et al., 2013, DOI 10.1161/ ATVBAHA.113.301981).
• ETC-216 also known as MDCO-216 is a lipid-depleted form of HDL containing recombinant ApoA-I Milano . See Nicholls et al., 2011, Expert Opin Biol Ther. 11(3):387-94. doi: 10.1517/14712598.2011.557061.
• a complex that can be used in the methods of the disclosure is CER-522.
• CER-522 is a lipoprotein complex comprising a combination of three phospholipids and a 22 amino acid peptide, CT80522:
• the phospholipid component of CER-522 consists 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 48.5:48.5:3 weight ratio.
• the ratio of peptide to total phospholipids in the CER-522 complex is 1:2.5 (w/w).
• the lipoprotein complex is delipidated HDL.
• Most HDL in plasma is cholesterol-rich.
• the lipids in HDL can be depleted, for example partially and/or selectively depleted, e.g., to reduce its cholesterol content.
• the delipidated HDL can resemble small ⁇ , pre ⁇ -1, and other pre ⁇ forms of HDL. A process for selective depletion of HDL is described in Sacks et al., 2009, J Lipid Res.50(5): 894–907.
• a lipoprotein complex comprises a bioactive agent delivery particle as described in US 2004/0229794.
• a bioactive agent delivery particle can comprise a lipid binding polypeptide (e.g., an apolipoprotein as described previously in this Section or in Section 6.1.2), a lipid bilayer (e.g., comprising one or more phospholipids as described previously in this Section or in Section 6.1.3.1), and a bioactive agent (e.g., an anti-cancer agent), wherein the interior of the lipid bilayer comprises a hydrophobic region, and wherein the bioactive agent is associated with the hydrophobic region of the lipid bilayer.
• a bioactive agent delivery particle as described in US 2004/0229794.
• a bioactive agent delivery particle does not comprise a hydrophilic core.
• a bioactive agent delivery particle is disc shaped (e.g., having a diameter from about 7 to about 29 nm).
• Bioactive agent delivery particles include bilayer-forming lipids, for example phospholipids (e.g., as described previously in this Section or in Section 6.1.3.1).
• a bioactive agent delivery particle includes both bilayer-forming and non-bilayer- forming lipids.
• the lipid bilayer of a bioactive agent delivery particle includes phospholipids.
• the phospholipids incorporated into a delivery particle include dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylglycerol (DMPG).
• the lipid bilayer includes DMPC and DMPG in a 7:3 molar ratio.
• the lipid binding polypeptide is an apolipoprotein (e.g., as described previously in this Section or in Section 6.1.2).
• the predominant interaction between lipid binding polypeptides, e.g., apolipoprotein molecules, and the lipid bilayer is generally a hydrophobic interaction between residues on a hydrophobic face of an amphipathic structure, e.g., an ⁇ -helix of the lipid binding polypeptide and fatty acyl chains of lipids on an exterior surface at the perimeter of the particle.
• Bioactive agent delivery particles may include exchangeable and/or non-exchangeable apolipoproteins.
• the lipid binding polypeptide is ApoA-I.
• bioactive agent delivery particles include lipid binding polypeptide molecules, e.g., apolipoprotein molecules, that have been modified to increase stability of the particle.
• the modification includes introduction of cysteine residues to form intramolecular and/or intermolecular disulfide bonds.
• bioactive agent delivery particles include a chimeric lipid binding polypeptide molecule, e.g., a chimeric apolipoprotein molecule, with one or more bound functional moieties, for example one or more targeting moieties and/or one or more moieties having a desired biological activity, e.g., antimicrobial activity, which may augment or work in synergy with the activity of a bioactive agent incorporated into the delivery particle. 6.1.2.
• Lipid binding protein molecules that can be used in the complexes described herein include apolipoproteins such as those described in Section 6.1.2.1 and apolipoprotein mimetic peptides such as those described in Section 6.1.2.2.
• the complex comprises a mixture of lipid binding protein molecules.
• the complex comprises a mixture of one or more lipid binding protein molecules and one or more apolipoprotein mimetic peptides.
• 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 terms of ApoA-I equivalents based upon the number of amphipathic helices they contain.
• ApoA-IM which typically exists as a disulfide-bridged dimer, can be expressed as 2 ApoA-I equivalents, because each molecule of ApoA-IM contains twice as many amphipathic helices as a molecule of ApoA-I.
• a peptide mimetic that contains a single amphipathic helix can be expressed as a 1/10-1/6 ApoA-I equivalent, because each molecule contains 1/10-1/6 as many amphipathic helices as a molecule of ApoA-I. 6.1.2.1.
• Suitable apolipoproteins that can be included in the lipid binding protein-based complexes include apolipoproteins ApoA-I, ApoA-II, ApoA-IV, ApoA-V, ApoB, ApoC-I, ApoC-II, ApoC-III, ApoD, ApoE, ApoJ, ApoH, and any combination of two or more of the foregoing.
• Apolipoprotein A-IMilano Apolipoprotein A-IMilano
• Apolipoprotein A-IParis Apolipoprotein A-IParis
• Apolipoprotein A-IZaragoza Apolipoprotein A-IZ
• Apolipoproteins mutants containing cysteine residues are also known, and can also be used (see, e.g., U.S. Publication No.2003/018132).
• the apolipoproteins may be in the form of monomers or dimers, which may be homodimers or heterodimers.
• apolipoproteins can be modified in their primary sequence to render them less susceptible to oxidations, for example, as described in U.S. Publication Nos.2008/0234192 and 2013/0137628, and U.S. Patent Nos.8,143,224 and 8,541,236.
• the apolipoproteins can include residues corresponding to elements that facilitate their isolation, such as His tags, or other elements designed for other purposes.
• 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).
• a biological fluid e.g., lymph, cerebrospinal fluid, vitreous humor, aqueous humor, blood, or a blood fraction (e.g., serum or plasma).
• the complex comprises covalently bound lipid-binding protein monomers, e.g., dimeric apolipoprotein A-IMilano, which is a mutated form of ApoA-I containing a cysteine.
• the cysteine allows the formation of a disulfide bridge which can lead to the formation of homodimers or heterodimers (e.g., ApoA-I Milano-ApoA-II).
• the apolipoprotein molecules comprise ApoA-I, ApoA-II, ApoA- IV, ApoA-V, ApoB, ApoC-I, ApoC-II, ApoC-III, ApoD, ApoE, ApoJ, or ApoH molecules or a combination thereof.
• the apolipoprotein molecules comprise or consist of ApoA-I molecules.
• said ApoA-I molecules are human ApoA-I molecules.
• said ApoA-I molecules are recombinant. In some embodiments, the ApoA- I molecules are not ApoA-IMilano. [0103] In some embodiments, the ApoA-I molecules are Apolipoprotein A-IMilano (ApoA-IM), Apolipoprotein A-IParis (ApoA-IP), or Apolipoprotein A-IZaragoza (ApoA-IZ) molecules. [0104] Apolipoproteins can be purified from animal sources (and in particular from human sources) or produced recombinantly as is well-known in the art, see, e.g., Chung et al., 1980, J.
• the apolipoprotein can be in prepro- form, pro- form, or mature form.
• a complex can comprise ApoA-I (e.g., human ApoA-I) in which the ApoA-I is preproApoA-I, proApoA-I, or mature ApoA-I.
• the complex comprises ApoA-I that has at least 90% sequence identity to SEQ ID NO:1: PPQSPWDRVKDLATVYVDVLKDSGRDYVSQFEGSALGKQLNLKLLDNWDSVTSTFSKLREQL GPVTQEFWDNLEKETEGLRQEMSKDLEEVKAKVQPYLDDFQKKWQEEMELYRQKVEPLRAE LQEGARQKLHELQEKLSPLGEEMRDRARAHVDALRTHLAPYSDELRQRLAARLEALKENGGA RLAEYHAKATEHLSTLSEKAKPALEDLRQGLLPVLESFKVSFLSALEEYTKKLNTQ (SEQ ID NO:1) [0106]
• SEQ ID NO:1 PP
• the complex comprises ApoA-I that has at least 98% sequence identity to SEQ ID NO:1. In other embodiments, the complex comprises ApoA-I that has at least 99% sequence identity to SEQ ID NO:1. In other embodiments, the complex comprises ApoA-I that has 100% sequence identity to SEQ ID NO:1. [0107] 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.
• 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.
• the apolipoprotein molecule(s) can comprise a chimeric apolipoprotein comprising an apolipoprotein and one or more attached functional moieties, such as for example, one or more CRN-001 complex(es), one or more targeting moieties, a moiety having a desired biological activity, an affinity tag to assist with purification, and/or a reporter molecule for characterization or localization studies.
• An attached moiety with biological activity may have an activity that is capable of augmenting and/or synergizing with the biological activity of a compound incorporated into a complex of the disclosure.
• a moiety with biological activity may have antimicrobial (for example, antifungal, antibacterial, anti-protozoal, bacteriostatic, fungistatic, or antiviral) activity.
• an attached functional moiety of a chimeric apolipoprotein is not in contact with hydrophobic surfaces of the complex.
• an attached functional moiety is in contact with hydrophobic surfaces of the complex.
• a functional moiety of a chimeric apolipoprotein may be intrinsic to a natural protein.
• a chimeric apolipoprotein includes a ligand or sequence recognized by or capable of interaction with a cell surface receptor or other cell surface moiety.
• a chimeric apolipoprotein includes a targeting moiety that is not intrinsic to the native apolipoprotein, such as for example, S. cerevisiae ⁇ -mating factor peptide, folic acid, transferrin, or lactoferrin.
• a chimeric apolipoprotein includes a moiety with a desired biological activity that augments and/or synergizes with the activity of a compound incorporated into a complex of the disclosure.
• a chimeric apolipoprotein may include a functional moiety intrinsic to an apolipoprotein.
• an apolipoprotein intrinsic functional moiety is the intrinsic targeting moiety formed approximately by amino acids 130-150 of human ApoE, which comprises the receptor binding region recognized by members of the low density lipoprotein receptor family.
• Other examples of apolipoprotein intrinsic functional moieties include the region of ApoB-100 that interacts with the low density lipoprotein receptor and the region of ApoA-I that interacts with scavenger receptor type B 1.
• a functional moiety may be added synthetically or recombinantly to produce a chimeric apolipoprotein.
• apolipoprotein with the prepro or pro sequence from another preproapolipoprotein (e.g., prepro sequence from preproapoA-II substituted for the prepro sequence of preproapoA-I).
• apolipoprotein for which some of the amphipathic sequence segments have been substituted by other amphipathic sequence segments from another apolipoprotein.
• chimeric refers to two or more molecules that are capable of existing separately and are joined together to form a single molecule having the desired functionality of all of its constituent molecules.
• the constituent molecules of a chimeric molecule may be joined synthetically by chemical conjugation or, where the constituent molecules are all polypeptides or analogs thereof, polynucleotides encoding the polypeptides may be fused together recombinantly such that a single continuous polypeptide is expressed.
• a chimeric molecule is termed a fusion protein.
• a "fusion protein” is a chimeric molecule in which the constituent molecules are all polypeptides and are attached (fused) to each other such that the chimeric molecule forms a continuous single chain.
• the various constituents can be directly attached to each other or can be coupled through one or more linkers.
• One or more segments of various constituents can be, for example, inserted in the sequence of an apolipoprotein, or, as another example, can be added N-terminal or C-terminal to the sequence of an apolipoprotein.
• a fusion protein can comprise an antibody light chain, an antibody fragment, a heavy-chain antibody, or a single-domain antibody.
• a chimeric apolipoprotein is prepared by chemically conjugating the apolipoprotein and the functional moiety to be attached. Means of chemically conjugating molecules are well known to those of skill in the art. Such means will vary according to the structure of the moiety to be attached, but will be readily ascertainable to those of skill in the art.
• Polypeptides typically contain a variety of functional groups, e.g., carboxylic acid (--COOH), free amino (--NH2), or sulfhydryl (--SH) groups, that are available for reaction with a suitable functional group on the functional moiety or on a linker to bind the moiety thereto.
• a functional moiety may be attached at the N-terminus, the C-terminus, or to a functional group on an interior residue (i.e., a residue at a position intermediate between the N- and C-termini) of an apolipoprotein molecule.
• the apolipoprotein and/or the moiety to be tagged can be derivatized to expose or attach additional reactive functional groups.
• fusion proteins that include a polypeptide functional moiety are synthesized using recombinant expression systems. Typically, this involves creating a nucleic acid (e.g., DNA) sequence that encodes the apolipoprotein and the functional moiety such 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.
• a nucleic acid encoding a chimeric apolipoprotein can be incorporated into a recombinant expression vector in a form suitable for expression in a host cell.
• an "expression vector” is a nucleic acid which, when introduced into an appropriate host cell, can be transcribed and translated into a polypeptide.
• the vector may also include regulatory sequences such as promoters, enhancers, or other expression control elements (e.g., polyadenylation signals).
• regulatory sequences are known to those skilled in the art (see, e.g., Goeddel, 1990, Gene Expression Technology: Meth.
• an apolipoprotein has been modified such that when the apolipoprotein is incorporated into a complex of the disclosure, the modification will increase stability of the complex, confer targeting ability or increase capacity.
• the modification includes introduction of cysteine residues into apolipoprotein molecules to permit formation of intramolecular or intermolecular disulfide bonds, e.g., by site-directed mutagenesis.
• a chemical crosslinking agent is used to form intermolecular links between apolipoprotein molecules to enhance stability of the complex. Intermolecular crosslinking prevents or reduces dissociation of apolipoprotein molecules from the complex and/or prevents displacement by endogenous apolipoprotein molecules within an individual to whom the complexes are administered.
• an apolipoprotein is modified either by chemical derivatization of one or more amino acid residues or by site directed mutagenesis, to confer targeting ability to or recognition by a cell surface receptor.
• Complexes can be targeted to a specific cell surface receptor by engineering receptor recognition properties into an apolipoprotein.
• complexes may be targeted to a particular cell type known to harbor a particular type of infectious agent, for example by modifying the apolipoprotein to render it capable of interacting with a receptor on the surface of the cell type being targeted.
• complexes may be targeted to macrophages by altering the apolipoprotein to confer recognition by the macrophage endocytic class A scavenger receptor (SR-A).
• SR-A macrophage endocytic class A scavenger receptor
• SR-A binding ability can be conferred to a complex by modifying the apolipoprotein by site directed mutagenesis to replace one or more positively charged amino acids with a neutral or negatively charged amino acid.
• SR-A recognition can also be conferred by preparing a chimeric apolipoprotein that includes an N- or C-terminal extension having a ligand recognized by SR-A or an amino acid sequence with a high concentration of negatively charged residues.
• Complexes comprising apoplipoproteins can also interact with apolipoprotein receptors such as, but not limited to, ABCA1 receptors, ABCG1 receptors, Megalin, Cubulin and HDL receptors such as SR-B1. 6.1.2.2.
• Apolipoprotein mimetics Peptides, peptide analogs, and agonists that mimic the activity of an apolipoprotein (collectively referred to herein as “apolipoprotein peptide mimetics”) can also be used in the complexes described herein, either alone, in combination with one or more other lipid binding proteins.
• apolipoprotein peptide mimetics can also be used in the complexes described herein, either alone, in combination with one or more other lipid binding proteins.
• Non-limiting examples of peptides and peptide analogs that correspond to apolipoproteins, as well as agonists that mimic the activity of ApoA-I, ApoA-IM, ApoA-II, ApoA- IV, and ApoE, that are suitable for inclusion in the complexes and compositions described herein are disclosed in U.S. Pat.
• WO/2010/093918 to Dasseux et al., the disclosures of which are incorporated herein by reference in their entireties.
• These peptides and peptide analogues can be composed of L-amino acid or D-amino acids or mixture of L- and D-amino acids. They may also include one or more non-peptide or amide linkages, such as one or more well-known peptide/amide isosteres.
• Such apolipoprotein peptide mimetic can be synthesized or manufactured using any technique for peptide synthesis known in the art, including, e.g., the techniques described in U.S. Pat. Nos.6,004,925, 6,037,323 and 6,046,166.
• the lipid binding protein molecules comprise apolipoprotein peptide mimetic molecules and optionally one or more apolipoprotein molecules such as those described above.
• the apolipoprotein peptide mimetic molecules comprise an ApoA-I peptide mimetic, ApoA-II peptide mimetic, ApoA-IV peptide mimetic, or ApoE peptide mimetic or a combination thereof.
• Amphipathic molecules [0119] An amphipathic molecule is a molecule that possesses both hydrophobic (apolar) and hydrophilic (polar) elements.
• Amphipathic molecules that can be used in complexes described herein include lipids (e.g., as described in Section 6.1.3.1), detergents (e.g., as described in Section 6.1.3.2), fatty acids (e.g., as described in Section 6.1.3.3), and apolar molecules and sterols covalently attached to polar molecules such as, but not limited to, sugars or nucleic acids (e.g., as described in Section 6.1.3.4).
• the complexes can include a single class of amphipathic molecule (e.g., a single species of phospholipids or a mixture of phospholipids) or can contain a combination of classes of amphipathic molecules (e.g., phospholipids and detergents).
• the complex can contain one species of amphipathic molecules or a combination of amphipathic molecules configured to facilitate solubilization of the lipid binding protein molecule(s).
• the amphipathic molecules included in comprise a phospholipid, a detergent, a fatty acid, an apolar moiety or sterol covalently attached to a sugar, or a combination thereof (e.g., selected from the types of amphipathic molecules discussed above).
• the amphipathic molecules comprise or consist of phospholipid molecules.
• the phospholipid molecules comprise negatively charged phospholipids, neutral phospholipids, positively charged phospholipids or a combination thereof.
• the phospholipid molecules contribute 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 molecules consist of a combination of negatively charged and neutral phospholipids. In some embodiments, the molar ratio of negatively charge phospholipid to neutral phospholipid ranges from 1:1 to 1:3. In some embodiments, the molar ratio of negatively charged phospholipid to neutral phospholipid is about 1:1 or about 1:2. [0123] In some embodiments, the amphipathic molecules comprise neutral phospholipids and negatively charged phospholipids in a weight ratio of 95:5 to 99:1. 6.1.3.1.
• Lipid binding protein-based complexes can include one or more lipids.
• one or more lipids can be saturated and/or unsaturated, natural and/or synthetic, charged or not charged, zwitterionic or not.
• the lipid molecules e.g., phospholipid molecules
• the lipid molecules can together contribute a net charge of 1-3 (e.g., 1-3, 1-2, 2-3, 1, 2, or 3) per lipid binding protein molecule in the complex.
• the net charge is negative.
• the net charge is positive.
• the lipid comprises a phospholipid.
• Phospholipids can have two acyl chains that are the same or different (for example, chains having a different number of carbon atoms, a different degree of saturation between the acyl chains, different branching of the acyl chains, or a combination thereof).
• the lipid can also be modified to contain a fluorescent probe (e.g., as described at yorkilipids.com/product-category/products/fluorescent- lipids/).
• the lipid comprises at least one phospholipid.
• 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).
• a phospholipid used in a complex of the disclosure has one or two acyl chains of 12, 14, 16, 18, 20, 22, or 24 carbons (e.g., two acyl chains of the same length or two acyl chains of different length).
• acyl chains present in commonly occurring fatty acids that can be included in phospholipids are provided in Table 1, below:
• Lipids that can be present in the complexes of the 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-
• Synthetic lipids such as synthetic palmitoylsphingomyelin or N-palmitoyl-4- hydroxysphinganine-1-phosphocholine (a form of phytosphingomyelin) can be used to minimize lipid oxidation.
• a lipid binding protein-based complex includes two types of phospholipids: a neutral lipid, e.g., lecithin and/or sphingomyelin (abbreviated SM), and a charged phospholipid (e.g., a negatively charged phospholipid).
• SM lecithin and/or sphingomyelin
• a “neutral” phospholipid has a net charge of about zero at physiological pH.
• neutral phospholipids are zwitterions, although other types of net neutral phospholipids are known and can be used.
• the molar ratio of the charged phospholipid (e.g., negatively charged phospholipid) to neutral phospholipid ranges from 1:1 to 1:3, for example, about 1:1, about 1:2, or about 1:3.
• the neutral phospholipid can comprise, for example, one or both of the lecithin and/or SM, and can optionally include other neutral phospholipids.
• the neutral phospholipid comprises lecithin, but not SM. In other embodiments, the neutral phospholipid comprises SM, but not lecithin.
• the neutral phospholipid comprises both lecithin and SM. All of these specific exemplary embodiments can include neutral phospholipids in addition to the lecithin and/or SM, but in many embodiments do not include such additional neutral phospholipids.
• SM includes sphingomyelins derived or obtained from natural sources, as well as analogs and derivatives of naturally occurring SMs that are impervious to hydrolysis by LCAT, as is naturally occurring SM.
• SM is a phospholipid very similar in structure to lecithin, but, unlike lecithin, it does not have a glycerol backbone, and hence does not have ester linkages attaching the acyl chains.
• SM has a ceramide backbone, with amide linkages connecting the acyl chains.
• SM can be obtained, for example, from milk, egg or brain.
• SM analogues or derivatives can also be used.
• Non-limiting examples of useful SM analogues and derivatives include, but are not limited to, palmitoylsphingomyelin, N-palmitoyl-4-hydroxysphinganine-1-phosphocholine (a form of phytosphingomyelin), palmitoylsphingomyelin, stearoylsphingomyelin, D-erythro-N-16:0-sphingomyelin and its dihydro isomer, D-erythro-N-16:0-dihydro-sphingomyelin.
• Synthetic SM such as synthetic palmitoylsphingomyelin or N-palmitoyl-4-hydroxysphinganine-1-phosphocholine (phytosphingomyelin) can be used in order to produce more homogeneous complexes and with fewer contaminants and/or oxidation products than sphingolipids of animal origin. Methods for synthesizing SM are described in U.S. Publication No.2016/0075634. [0132] Sphingomyelins isolated from natural sources can be artificially enriched in one particular saturated or unsaturated acyl chain.
• milk sphingomyelin (Avanti Phospholipid, Alabaster, Ala.) is characterized by long saturated acyl chains (i.e., acyl chains having 20 or more carbon atoms).
• egg sphingomyelin is characterized by short saturated acyl chains (i.e., acyl chains having fewer than 20 carbon atoms).
• milk sphingomyelin comprises C16:0 (16 carbon, saturated) acyl chains
• egg sphingomyelin comprises C16:0 acyl chains.
• the composition of milk sphingomyelin can be enriched to have an acyl chain composition comparable to that of egg sphingomyelin, or vice versa.
• the SM can be semi-synthetic such that it has particular acyl chains.
• milk sphingomyelin can be first purified from milk, then one particular acyl chain, e.g., the C16:0 acyl chain, can be cleaved and replaced by another acyl chain.
• the SM can also be entirely synthesized, by e.g., large-scale synthesis. See, e.g., Dong et al., U.S. Pat.
• SM can be fully synthetic, e.g., as described in U.S. Publication No. 2014/0275590.
• the lengths and saturation levels of the acyl chains comprising a semi-synthetic or a synthetic SM can be selectively varied.
• the acyl chains can be saturated or unsaturated, and can contain from about 6 to about 24 carbon atoms. Each chain can contain the same number of carbon atoms or, alternatively each chain can contain different numbers of carbon atoms.
• the semi-synthetic or synthetic SM comprises mixed acyl chains such that one chain is saturated and one chain is unsaturated.
• the chain lengths can be the same or different.
• the acyl chains of the semi- synthetic or synthetic SM are either both saturated or both unsaturated. Again, the chains can contain the same or different numbers of carbon atoms.
• both acyl chains comprising the semi-synthetic or synthetic SM are identical.
• the chains correspond to the acyl chains of a naturally-occurring fatty acid, such as for example oleic, palmitic or stearic acid.
• SM with saturated or unsaturated functionalized chains is used.
• both acyl chains are saturated and contain from 6 to 24 carbon atoms.
• Non-limiting examples of acyl chains present in commonly occurring fatty acids that can be included in semi-synthetic and synthetic SMs are provided in Table 1, above.
• the SM is palmitoyl SM, such as synthetic palmitoyl SM, which has C16:0 acyl chains, or is egg SM, which includes as a principal component palmitoyl SM.
• functionalized SM such as phytosphingomyelin, is used.
• Lecithin can be derived or isolated from natural sources, or it can be obtained synthetically.
• suitable lecithins isolated from natural sources include, but are not limited to, egg phosphatidylcholine and soybean phosphatidylcholine.
• lecithins include, dipalmitoylphosphatidylcholine, dimyristoylphosphatidylcholine, distearoylphosphatidylcholine 1-myristoy1-2- palmitoylphosphatidylcholine, 1-palmitoy1-2-myristoylphosphatidylcholine, 1-palmitoy1-2- stearoylphosphatidylcholine, 1-stearoy1-2-palmitoylphosphatidylcholine, 1-palmitoy1-2- oleoylphosphatidylcholine, 1-oleoy1-2-palmitylphosphatidylcholine, dioleoylphosphatidylcholine and the ether derivatives or analogs thereof.
• Lecithins derived or isolated from natural sources can be enriched to include specified acyl chains.
• identity(ies) of the acyl chains can be selectively varied, as discussed above in connection with SM.
• both acyl chains on the lecithin are identical.
• the acyl chains of the SM and lecithin are all identical.
• the acyl chains correspond to the acyl chains of myristitic, palmitic, oleic or stearic acid.
• the complexes of the disclosure can include one or more negatively charged phospholipids (e.g., alone or in combination with one or more neutral phospholipids).
• negatively charged phospholipids are phospholipids that have a net negative charge at physiological pH.
• the negatively charged phospholipid can comprise a single type of negatively charged phospholipid, or a mixture of two or more different, negatively charged, phospholipids.
• the charged phospholipids are negatively charged glycerophospholipids.
• Suitable negatively charged phospholipids include, but are not limited to, a 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)], a phosphatidylglycerol, a phospatidylinositol, a phosphatidylserine, a phosphatidic acid, and salts thereof (e.g., sodium salts or potassium salts).
• the negatively charged phospholipid comprises one or more of phosphatidylinositol, phosphatidylserine, phosphatidylglycerol and/or phosphatidic acid.
• the negatively charged phospholipid comprises or consists of a salt of a phosphatidylglycerol or a salt of a phosphatidylinositol.
• the negatively charged phospholipid comprises or consists of 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)], or DPPG, or a salt thereof.
• the negatively charged phospholipids can be obtained from natural sources or prepared by chemical synthesis. In embodiments employing synthetic negatively charged phospholipids, the identities of the acyl chains can be selectively varied, as discussed above in connection with SM.
• both acyl chains on the negatively charged phospholipids are identical.
• the acyl chains all types of phospholipids included in a complex of the disclosure are all identical.
• the complex comprises negatively charged phospholipid(s), and/or SM all having C16:0 or C16:1 acyl chains.
• the fatty acid moiety of the SM is predominantly C16:1 palmitoyl.
• the acyl chains of the charged phospholipid(s), lecithin and/or SM correspond to the acyl chain of palmitic acid.
• the acyl chains of the charged phospholipid(s), lecithin and/or SM correspond to the acyl chain of oleic acid.
• positively charged phospholipids that can be included in the complexes of the disclosure include N1-[2-((1S)-1-[(3-aminopropyl)amino]-4-[di(3-amino- propyl)amino]butylcarboxamido)ethyl]-3,4-di[oleyloxy]-benzamide, 1,2-di-O-octadecenyl-3- trimethylammonium propane, 1,2-dimyristoleoyl-sn-glycero-3-ethylphosphocholine, 1-palmitoyl- 2-oleoyl-sn-glycero-3-ethylphosphocholine, 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine
• the lipids used are preferably at least 95% pure, and/or have reduced levels of oxidative agents (such as but not limited to peroxides).
• Lipids obtained from natural sources preferably have fewer polyunsaturated fatty acid moieties and/or fatty acid moieties that are not susceptible to oxidation.
• the level of oxidation in a sample can be determined using an iodometric method, which provides a peroxide value, expressed in milli-equivalent number of isolated iodines per kg of sample, abbreviated meq O/kg.
• the level of oxidation, 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.
• Complexes can in some embodiments include small quantities of additional lipids.
• lipids can be used, including, but not limited to, lysophospholipids, galactocerebroside, gangliosides, cerebrosides, glycerides, triglycerides, and sterols and sterol derivatives (e.g., a plant sterol, an animal sterol, such as cholesterol, or a sterol derivative, such as a cholesterol derivative).
• a complex of the disclosure can contain cholesterol or a cholesterol derivative, e.g., a cholesterol ester.
• the cholesterol derivative can also be a substituted cholesterol or a substituted cholesterol ester.
• the complexes of the disclosure can also contain an oxidized sterol such as, but not limited to, oxidized cholesterol or an oxidized sterol derivative (such as, but not limited to, an oxidized cholesterol ester). In some embodiments, the complexes do not include cholesterol and/or its derivatives (such as a cholesterol ester or an oxidized cholesterol ester).
• oxidized sterol such as, but not limited to, oxidized cholesterol or an oxidized sterol derivative (such as, but not limited to, an oxidized cholesterol ester).
• the complexes do not include cholesterol and/or its derivatives (such as a cholesterol ester or an oxidized cholesterol ester).
• the complexes can contain one or more detergents.
• the detergent can be zwitterionic, nonionic, cationic, anionic, or a combination thereof.
• Exemplary zwitterionic detergents include 3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), 3-[(3- Cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonate (CHAPSO), and N,N- dimethyldodecylamine N-oxide (LDAO).
• CHAO 3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate
• CHAO N,N- dimethyldodecylamine N-oxide
• nonionic detergents include D-(+)-trehalose 6-monooleate, N-octanoyl-N-methylglucamine, N-nonanoyl-N-methylglucamine, N-decanoyl-N- methylglucamine, 1-(7Z-hexadecenoyl)-rac-glycerol, 1-(8Z-hexadecenoyl)-rac-glycerol, 1-(8Z- heptadecenoyl)-rac-glycerol, 1-(9Z-hexadecenoyl)-rac-glycerol, 1-decanoyl-rac-glycerol.
• Exemplary cationic detergents include (S)-O-methyl-serine dodecylamide hydrochloride, dodecylammonium chloride, decyltrimethylammonium bromide, and cetyltrimethylammonium sulfate.
• Exemplary anionic detergents include cholesteryl hemisuccinate, cholate, alkyl sulfates, and alkyl sulfonates.
• 6.1.3.3. Fatty Acids [0145]
• the complexes can contain one or more fatty acids.
• the one or more fatty acids can include short-chain fatty acids having aliphatic tails of five or fewer carbons (e.g.
• butyric acid, isobutyric acid, valeric acid, or isovaleric acid medium-chain fatty acids having aliphatic tails of 6 to 12 carbons (e.g., caproic acid, caprylic acid, capric acid, or lauric acid), long-chain fatty acids having aliphatic tails of 13 to 21 carbons (e.g., myristic acid, palmitic acid, stearic acid, or arachidic acid) , very long chain fatty acids having aliphatic tails of 22 or more carbons (e.g., behenic acid, lignoceric acid, or cerotic acid), or a combination thereof.
• medium-chain fatty acids having aliphatic tails of 6 to 12 carbons e.g., caproic acid, caprylic acid, capric acid, or lauric acid
• long-chain fatty acids having aliphatic tails of 13 to 21 carbons e.g., myristic acid, palmitic acid, stearic acid
• 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, sapienic acid, oleic acid, elaidic acid, vaccenic acid, linoleic acid, linoelaidic acid, ⁇ -linolenic acid, arachidonic acid, eicosapentaenoic acid, erucic acid, or docosahexaenoic acid) or a combination thereof.
• 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
• Unsaturated fatty acids can be cis or trans fatty acids.
• unsaturated fatty acids used in the complexes of the disclosure are cis fatty acids.
• the complexes can contain one or more amphipathic molecules that comprise an apolar molecule or moiety (e.g., a hydrocarbon chain, an acyl or diacyl chain) or a sterol (e.g., cholesterol) attached to a sugar (e.g., a monosaccharide such as glucose or galactose, or a disaccharide such as maltose or trehalose).
• the sugar can be a modified sugar or a substituted sugar.
• Exemplary amphipathic molecules comprising an apolar molecule attached to a sugar include dodecan-2-yloxy-ß-D-maltoside, tridecan-3-yloxy-ß-D-maltoside, tridecan-2-yloxy-ß-D- maltoside, n-dodecyl-ß-D-maltoside (DDM), n-octyl-ß-D-glucoside, n-nonyl-ß-D-glucoside, n- decyl-ß-D-maltoside, n-dodecyl- ⁇ -D-maltopyranoside, 4-n-Dodecyl- ⁇ , ⁇ trehalose, 6-n-dodecyl- ⁇ , ⁇ trehalose, and 3-n-dodecyl- ⁇ , ⁇ trehalose.
• DDM dodecan-2-yloxy-ß-D-maltoside
• tridecan-3-yloxy-ß-D-maltoside tri
• the apolar moiety is an acyl or a diacyl chain.
• the sugar is a modified sugar or a substituted sugar. 6.1.4.
• Formulations [0149] Lipid binding protein-based complexes can be formulated for 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). [0150] CER-001 intended for administration by infusion can be formulated in a phosphate buffer with sucrose and mannitol excipients, for example as described in WO 2012/109162. 6.2.
• Subjects who can be treated according to the methods described herein are preferably mammals, most preferably human.
• the subject has an acute condition comprising acute inflammation.
• the subject can be a subject in need of therapy for sepsis and/or AKI.
• the subject has sepsis (e.g., associated with a gram-negative bacterial infection).
• the sepsis can in some embodiments be caused by an intra-abdominal cavity infection or be urosepsis. Sepsis is a risk factor for AKI.
• the subject can be at risk for AKI, for example due to sepsis.
• the subject has sepsis associated with a gram negative bacterial infection. In other embodiments, the subject has sepsis associated with a gram positive bacterial infection. [0155] In some embodiments, the subject has a SOFA score of 1 to 4 before treatment with a lipid binding protein-based complex, e.g., a score of 1, 2, 3, or 4 (see, Vincent et al.1996, Intensive Care Med, 22:707–710).
• the subject has an endotoxin activity level as measured by the Endotoxin Activity Assay (EEATM) (Spectral Medical) of > 0.6 prior to administration of the lipid binding protein-based complex (see, Marshall et al., 2004, J Infect Dis.190(3):527-34).
• EAATM Endotoxin Activity Assay
• the subject has AKI or is at risk of AKI.
• a the AKI can be sepsis-related AKI, ischemia/reperfusion AKI, CSA-AKI, or hepatorenal syndrome (HSA) AKI.
• the AKI is sepsis-related AKI.
• the AKI is ischemia/reperfusion AKI.
• the AKI is CSA AKI. In other embodiments, the AKI is HRS AKI.
• Subjects at risk of HRS include subjects having liver disease (e.g., chronic liver disease or acute liver disease). In some embodiments, the subject has chronic liver disease. In some embodiments, the subject has acute liver disease. In some embodiments, the subject has alcoholic liver disease. HRS has historically been classified as type 1 HRS, where renal function rapidly deteriorates over days to weeks, and type 2 HRS, where deterioration occurs over months. Accordingly, in some embodiments, a subject treated according to a dosage regimen of the disclosure has type 1 HRS. In other embodiments, a subject treated according to a dosage regiment of the disclosure has type 2 HRS.
• a subject having HRS meets the ICA diagnostic criteria of HRS AKI.
• the subject can be any subject having CRS or at risk of CRS, and/or any subject in need of reduction in serum levels of one or more inflammatory markers such as IL-6.
• the subject has CRS.
• the subject has CRS secondary to an infection, for example a viral infection such as an infection with COVID-19 or influenza.
• the subject has CRS secondary to a COVID-19 infection.
• the subject has CRS caused by immunotherapy, for example antibody or chimeric antigen receptor (CAR) T cell therapy.
• CAR chimeric antigen receptor
• the subject is at risk of CRS, for example due to an infection such as COVID-19 or influenza.
• the subject is at risk of CRS due to immunotherapy.
• the subject is a subject in need of a reduction in serum levels of one or more inflammatory markers, for example a subject with elevated levels of the 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 ⁇ (TNF ⁇ ).
• the one or more cytokines comprise IL-6.
• the one or more cytokines comprise a combination of the foregoing, for example, 2, 3, 4, 5, 6, 7, or all 8 of interleukin 6 (IL-6), C-reactive protein, D- dimer, ferritin, interleukin 8 (IL-8), granulocyte-macrophage colony stimulating factor (GM-CSF), monocyte chemoattractant protein (MCP) 1, and tumor necrosis factor ⁇ (TNF ⁇ ).
• IL-6 interleukin 6
• C-reactive protein D- dimer
• ferritin interleukin 8
• GM-CSF granulocyte-macrophage colony stimulating factor
• MCP monocyte chemoattractant protein
• TNF ⁇ tumor necrosis factor ⁇
• an administration regimen can include four or more doses of a lipid binding protein-based complex (e.g.,CER-001), e.g., five, six, seven, eight, nine, ten, eleven, twelve, or more than twelve doses.
• a lipid binding protein-based complex e.g.,CER-001
• the lipid binding protein-based complex is administered according to an induction and, optionally, a consolidation regimen as described in Sections 6.3.1 and 6.3.2, respectively.
• the lipid binding protein-based complex can be administered in a single phase, e.g., according to an administration regimen described in this Section.
• the subject is not treated with the lipid binding protein- based complex according to a maintenance regimen, e.g., a regimen comprising long-term (e.g., one month or longer) administration of the lipid binding protein-based complex.
• a maintenance regimen e.g., a regimen comprising long-term (e.g., one month or longer) administration of the lipid binding protein-based complex.
• the lipid binding protein-based complex (e.g., CER-001) administration regimens of the disclosure can last up to one week, one week, or more than one week (e.g., two weeks).
• a lipid binding protein-based complex (e.g., CER-001) administration regimen can comprise administering: - five doses of CER-001 over one week; - six doses of CER-001 over one week; - seven doses of CER-001 over one week; - ten doses of CER-001 over two weeks; - twelve doses of CER-001 over two weeks; - fourteen doses of CER-001 over two weeks.
• the methods of the disclosure e.g., methods for treating CRS or a subject at risk of CRS
• a lipid binding protein-based complex (e.g., CER-001) is administered daily, e.g., daily for at least 5 days, at least 6 days, at least 7 days, or more than 7 days (e.g., daily for up to one week or daily for up to two weeks).
• a lipid binding protein-based complex (e.g., CER-001) is administered less frequently, e.g., every other day, two times per week, three times per week, or once a week.
• an administration window can be provided, for example, to accommodate slight variations to a multi-dosing per week dosing schedule.
• a lipid binding protein-based complex (e.g., CER-001) can be administered in the methods of the disclosure for a pre-determined period of time, e.g., for one week.
• a lipid binding protein-based complex e.g., CER-001
• administration of a lipid binding protein-based complex can be continued until one or more symptoms of the acute indication (e.g., CRS) are reduced or continued until the serum levels of one or more inflammatory markers are reduced, for example reduced to a normal level or reduced relative to a baseline value for the subject, e.g., a baseline value measured prior to the start of lipid binding protein-based complex (e.g., CER-001) therapy.
• Reference or “normal” levels of various inflammatory markers are known in the art.
• the Mayo Clinic Laboratories 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 (males), 11-307 mcg/L (females); IL-8 ⁇ 57.8 pg/mL; TNF- ⁇ ⁇ 5.6 pg/mL.
• FEU Fibrinogen Equivalent Units
• a lipid binding protein-based complex e.g., CER-001
• a lipid binding protein-based complex can be administered before the immunotherapy begins, concurrently with the immunotherapy, after the immunotherapy ends, or a combination thereof.
• a lipid binding protein-based complex e.g., CER-001
• Concurrent administration is not limited to administration of the lipid binding protein-based complex (e.g., CER-001) and the immunotherapy at the exact same time, and encompasses administration of one agent while a course of therapy with the other is ongoing.
• the methods of the disclosure typically comprise administering a high dose of a lipid binding protein-based complex (e.g., CER-001).
• the high dose can be the aggregate of multiple individual doses (e.g., two, three, four, five, six, seven, eight, nine or 10 individual doses), for example administered over multiple days (e.g., a period of three days, four days, five days, six days, seven days, eight days, nine days, 10 days, eleven days, 12 days, 13 days, 14 days or 15 days).
• the individual doses of a high dose are in some embodiments administered daily, twice daily, or two to three days apart.
• 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, e.g., measured by circulating vascular cell adhesion molecule 1 (VCAM-1) and/or intercellular adhesion molecule 1 (ICAM-1) levels.
• VCAM-1 circulating vascular cell adhesion molecule 1
• IAM-1 intercellular adhesion molecule 1
• the high dose or an individual dose is an amount which 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.
• the high dose is an amount effective to reduce serum levels of one or more inflammatory markers, for example, one or more of IL-6, C-reactive protein, D- dimer, ferritin, IL-8, GM-CSF, and MCP1 TNF- ⁇ .
• the serum levels of the one or more inflammatory markers are reduced from an elevated range to a normal range, and/or reduced by at least 20%, at least 40%, or at least 60%.
• the dose of a lipid binding protein-based complex (e.g., CER-001) administered to a subject can in some embodiments range from 4 to 40 mg/kg (e.g., 10 to 40 mg/kg) on a protein weight basis (e.g., 5, 10, 15, 20, 25, 30, 35, or 40 mg/kg or any range bounded 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).
• protein weight basis means that a dose of a lipid binding protein-based complex (e.g., CER-001) to be administered to a subject is calculated based upon the amount of ApoA-I in the lipid binding protein-based complex (e.g., CER-001) to be administered and the weight of the subject. For example, a subject who weighs 70 kg and is to receive a 20 mg/kg dose of CER-001 would receive an amount of CER-001 that provides 1400 mg of ApoA-I (70 kg x 20 mg/kg).
• a lipid binding protein-based complex e.g., CER-001
• the unit dosage used in the methods of the disclosure can in some embodiments vary from 300 mg to 4000 mg (e.g., 600 mg to 4000 mg) per administration (on a protein weight basis).
• the dosage of a lipid binding protein-based complex e.g., CER-001
• the dosage of a lipid binding protein-based complex is 600 mg to 3000 mg, 800 mg to 3000 mg, 1000 mg to 2400 mg, or 1000 mg to 2000 mg per administration (on a protein weight basis).
• a high dose of a lipid binding protein-based complex e.g., CER-001
• the aggregate of multiple individual doses is 600 mg to 40 g (on a protein weight basis).
• a high dose is 3 g to 35 g or 5 g to 30 g (on a protein weight basis).
• a lipid binding protein-based complex e.g., CER-001
• CER-001 is preferably administered as an IV infusion.
• a stock solution of CER-001 can be diluted in normal saline such as physiological saline (0.9% NaCl) to a total volume between 125 and 250 ml.
• subjects weighing less than 80 kg will have a total volume of 125 ml whereas subjects weighing at least 80 kg will have a total volume of 250 ml.
• doses of CER-001 are administered in a total volume of 250 ml.
• a lipid binding protein-based complex may be administered over a period ranging from one-hour to 24- hours. Depending on the needs of the subject, administration can be by slow infusion with a duration of more than one hour (e.g., up to 2 hours or up to 24 hours), by rapid infusion of one hour or less, or by a single bolus injection.
• a lipid binding protein-based complex e.g., CER-001
• is administered over a one-hour period e.g., using an infusion pump at a fixed rate of 125 ml/hr or 250 ml/hr.
• a dose of a lipid binding protein- based complex is administered as an infusion over a 24-hour period. 6.3.1. Induction Regimen [0177]
• induction regimens suitable for use in the methods of the disclosure entail administering multiple doses of a lipid binding protein-based complex (e.g., CER-001) over multiple consecutive days, e.g., three consecutive days.
• induction regimens suitable for use in the methods of the disclosure entail twice daily administration of a lipid binding protein-based complex (e.g., CER- 001) such as twice daily administration on multiple consecutive days.
• Twice daily administration can comprise, for example, two doses approximately 12 hours apart or a morning dose and an evening dose (which may be more or less than 12 hours apart).
• the induction regimen comprises two doses of a lipid binding protein- based complex (e.g., CER-001) per day for 3 consecutive days.
• a therapeutic dose of a lipid binding protein-based complex (e.g., CER-001) administered by infusion in the induction regimen can range from 4 to 40 mg/kg (e.g., 4 to 30 mg/kg) on a protein weight basis (e.g., 4, 5, 6, 7, 8, 9, 10, 1215, 20, 25, 30 or 40 mg/kg, or any range bounded 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).
• the dose of a lipid binding protein-based complex (e.g., CER-001) used in the induction regimen is 5 mg/kg.
• the dose of a lipid binding protein-based complex (e.g., CER-001) used in the induction regimen is 10 mg/kg. In some embodiments, the dose of a lipid binding protein-based complex (e.g., CER-001) used in the induction regimen is 15 mg/kg. In some embodiments, the dose of a 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 six doses of a lipid binding protein-based complex (e.g., CER-001) administered over three days at a dose of 5 mg/kg, 10 mg/kg, 15 mg/kg or 20 mg/kg.
• a lipid binding protein-based complex e.g., CER-001
• a lipid binding protein-based complex (e.g., CER-001) can be administered on a unit dosage basis.
• the unit dosage used in the induction phase can vary from 300 mg to 4000 mg (e.g., 300 mg to 3000 mg) (on a protein weight basis) per administration by infusion.
• the dosage of a lipid binding protein-based complex (e.g., CER-001) used during the induction phase is 300 mg to 1500 mg, 400 mg to 1500 mg, 500 mg to 1200 mg, or 500 mg to 1000 mg (on a protein weight basis) per administration by infusion. 6.3.2.
• Consolidation regimens suitable for use in the methods of the disclosure entail administering one dose or multiple doses of a lipid binding protein-based complex (e.g., CER- 001) following an induction regimen.
• the consolidation regimen comprises administering two doses of a lipid binding protein-based complex (e.g., CER-001).
• the two doses can be administered approximately 12 hours apart, or administered as a morning dose and an evening dose (which may be more or less than 12 hours apart).
• the dose(s) of a lipid binding protein-based complex (e.g., CER-001) in a consolidation regimen can in some embodiments be administered on day 6 of a dosing regimen that begins with an induction regimen on day 1.
• the dose(s) of a lipid binding protein-based complex (e.g., CER-001) in a consolidation regimen can in some embodiments be administered on day 4 of a dosing regimen that begins with an induction regimen on day 1.
• the dose(s) of a lipid binding protein-based complex (e.g., CER-001) in a consolidation regimen can in some embodiments be administered on day 5 of a dosing regimen that begins with an induction regimen on day 1.
• the dose(s) of a lipid binding protein-based complex (e.g., CER-001) in a consolidation regimen can in some embodiments be administered on day 7 of a dosing regimen that begins with an induction regimen on day 1.
• a therapeutic dose of a lipid binding protein-based complex (e.g., CER-001) administered by infusion in the consolidation regimen can range from 4 mg/kg to 40 mg/kg (e.g., 4 to 30 mg/kg) on a protein weight basis (e.g., 4, 5, 6, 7, 8, 9, 10, 12, 15, 20, 25, 30, or 40 mg/kg, or any range bounded 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).
• the dose of a lipid binding protein-based complex (e.g., CER-001) used in the consolidation regimen is 5 mg/kg. In some embodiments, the dose of a lipid binding protein-based complex (e.g., CER-001) used in the consolidation regimen is 10 mg/kg. In some embodiments, the dose of a lipid binding protein-based complex (e.g., CER-001) in the consolidation regimen is 15 mg/kg. In some embodiments, the dose of a lipid binding protein-based complex (e.g., CER-001) used in the consolidation regimen is 20 mg/kg.
• the consolidation regimen comprises two doses of a lipid binding protein-based complex (e.g., CER-001) administered on one day at a dose of 5 mg/kg, 10 mg/kg, 15 mg/kg or 20 mg/kg.
• a lipid binding protein-based complex e.g., CER-001
• the unit dosage used in the consolidation phase can vary from 300 mg to 4000 mg (e.g., 300 mg to 3000 mg) (on a protein weight basis) per administration by infusion.
• the dosage of a lipid binding protein-based complex (e.g., CER-001) used during the consolidation phase is 300 mg to 1500 mg, 400 mg to 1500 mg, 500 mg to 1200 mg, or 500 mg to 1000 mg (on a protein weight basis) per administration by infusion.
• the lipid binding protein-based complex (e.g., CER-001) can be administered during the consolidation phase in the same manner as described in Section 6.3, e.g., as an IV infusion over a one-hour period. 6.4.
• a lipid binding protein-based complex (e.g., CER-001) can be administered to a subject as described herein as a monotherapy or a part of a combination therapy regimen.
• a combination therapy may comprise a lipid binding protein-based complex (e.g., CER-001) in combination with a standard of care treatment for sepsis and/or AKI. See, e.g., Rhodes et al., 2017, Intensive Care Med 43:304–377; Dugar et al., 2020, Cleveland Clinic Journal of Medicine 87(1):53-64.
• the subject is treated with a lipid binding protein-based complex (e.g., CER-001) in combination with fluid replacement therapy.
• a lipid binding protein-based complex e.g., CER-001
• an antimicrobial e.g., antimicrobial-derived antimicrobial-derived antimicrobial-derived antimicrobial-derived antimicrobial-based complex
• the 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).
• an antibiotic e.g., ceftriaxone, meropenem, ceftazidime, cefotaxime, cefepime, piperacillin and tazobactam, ampicillin and sulbactam, imipenem and cilastatin, levofloxacin, or clindamycin.
• the subject is treated with a lipid binding protein-based complex (e.g., CER-001) in combination with an antiviral.
• the subject is treated with a lipid binding protein-based complex (e.g., CER-001) in combination with a medication that raises blood pressure (e.g., norepinephrine or epinephrine).
• a combination therapy regimen can in some embodiments comprise 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, olokizumab, elsilimomab, BMS-945429, sirukumab, levilimab, and CPSI-2364.
• a lipid binding protein-based complex e.g., CER-001 is administered in combination with tocilizumab.
• Subjects who have or have had a 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 spike protein of COVID-19, one or more antiviral agents (e.g., lopinavir, remdesivir, danoprevir, galidesivir, darunavir, ritonavir), chloroquine, hydroxychloroquine, azithromycin, an interferon (e.g., an interferon alpha or an interferon beta, each of which can be pegylated), or a combination thereof.
• a lipid binding protein-based complex e.g., CER-001
• additional therapies such as antibodies from recovered COVID-19 patients, antibodies against the spike protein of COVID-19, one or more antiviral agents (e.g., lopinavir, remdesivir, danoprevir, gal
• an antihistamine e.g., diphenhydramine, cetirizine, fexofenadine, or loratadine
• a lipid binding protein- based complex e.g., CER-001
• the antihistamine can reduce the likelihood of allergic reactions.
• Example 1 CER-001 therapy in a swine model of LPS-induced AKI
• LPS lipopolysaccharide
• Sepsis was induced in the pigs by intravenous infusion of a saline solution containing 300 ⁇ g/kg of LPS at T0.
• CER-001 treated pigs and CER-001 multiple dose treated pigs received a 20 mg/kg dose of CER-001 at T0.
• CER-001 multiple dose treated pigs 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. 7.1.2. Results [0197] An increased survival rate was observed in both CER-001 treated groups compared to LPS group (data not shown). LPS injection led to a time-dependent increase of IL-6 in endotoxemic animals (FIG.1) compared to the basal condition (T0).
• CER-001 treatment was able to reverse LPS effects, as shown by reduced IL-6 levels (FIG.1, “20 MG” and “40MG”).
• the second infusion of CER-001 three hours from the first dose (T3) strongly reduced IL-6 serum levels to basal level by the end of the study (T end) (FIG.1, “40MG”).
• T end the end of the study
• high levels of MCP-1 in endotoxemic pigs were observed relative to the basal condition, while MCP- 1 levels were lower in the pigs treated with CER-001 (data not shown).
• Endothelial dysfunction was evaluated by measuring sVCAM-1 and sICAM-1 serum levels.
• Example 2 Randomized pilot study comparing short-term CER-001 infusions at different doses to prevent sepsis-induced acute kidney injury [0201]
• SOC standard of care
• Study population This is a single-center, randomized, dose-ranging (phase II) study including patients with sepsis due to intra-abdominal cavity infection or urosepsis, admitted at the Intensive Care Unit (ICU) of the participating center. The investigators ensure that all patients meeting the following inclusion and exclusion criteria are offered enrollment in the study.
• ICU Intensive Care Unit
• Inclusion criteria - Male or non-pregnant female adult ⁇ 18 years of age at time of enrollment; - Meets Sepsis 3 criteria, defined as an acute increase of at least 2 points in SOFA Score relative to the SOFA score upon admission; - Endotoxin level (measured by Endotoxin Activity Assay (EEATM); Spectral Medical) >0.6 (see, Marshall et al., 2004, J Infect Dis.190(3):527-34); - Signed and dated informed consent by the patient itself or by a legal representative.
• Exclusion Criteria - Patients weighing more than 100 kg; - Alanine transaminase/aspartate transaminase (ALT/AST) > 5 times the upper limit of normal; - Stage 4 severe chronic kidney disease or requiring dialysis (i.e.
• estimated glomerular filtration rate ⁇ 30 ml /min/1.73 m 2 ); - Leukocytes ⁇ 2.0 ⁇ 10 ⁇ 9; - Pregnancy or breast feeding; - Undergone organ transplantation during the past one year; - Anticipated transfer to another hospital, which is not a study site within 72 hours; - Terminally ill, including metastases or hematological malignancy, with a life expectancy less than 30 days (as assessed by the attending physician) or have been classified as "Do Not Resuscitate"; - Previous history of end stage chronic organ failure(s); - Diagnosed with HIV; - Uncontrolled hemorrhage within the last 24 h; - Patients who have used an investigational drug or device within 30 days of the first dose of CER-001.
• eGFR estimated glomerular filtration rate
• Secondary endpoint Secondary endpoints are: - Change in endotoxin and IL-6 levels from baseline to Day 3, Day 6 and Day 9. - Baseline is defined as the last measurements taken prior to dosing on Day 1. - Change in the SOFA score (Vincent et al.1996, Intensive Care Med, 22:707–710) from baseline to Day 3, Day 6 and Day 9.
• Intervention/exposure Twenty patients meeting the eligibility criteria, who sign and date an ethical committee (EC)-approved informed consent form, are randomized and assigned (1:1:1:1) ratio to conventional therapy (Group A), low dose CER-001 (Group B) or medium dose CER-001 (Group C) or high dose CER-001 (Group D). Conventional therapy is modulated according to the clinical conditions. All non-experimental treatments are allowed to be administered concomitantly during the patient’s participation in this study: any medication the patient takes, other than study drugs specified per protocol, is considered a concomitant medication and is recorded in the study records. [0210] Each patient is identified at the screening by a patient number. Once assigned to a patient, the patient number is not reused.
• Treatment group All patients receive conventional therapy. Treated groups receive an additional therapy with the study drugs.
• - Group A Conventional therapy (i.e., antibiotic treatments and hemodynamic support according to patient’s conditions).
• - Group B Conventional therapy + CER-0015 mg/kg BID for 3 consecutive days, followed by 5 mg/kg BID on Day 6.
• - Group C Conventional therapy + CER-00110 mg/kg BID for 3 consecutive days, followed by 10 mg/kg BID on Day 6.
• Group D Conventional therapy + CER-00120 mg/kg BID for 3 consecutive days, followed by 20 mg/kg BID on Day 6.
• Patients are pretreated with antihistamine prior to each CER-001 dose (e.g. dexchlorpheniramine 5 mg or hydroxyzine 100 mg) to avoid any potential infusion reactions. Patients may be interrupted or discontinued from study medication if any of the following occur: [0213] Any drug-related adverse event or other reason which, in the Investigator's opinion, jeopardizes the patient's participation in the trial or the interpretation of trial data (e.g., severe inter-current illness requiring additional care measures or preventing further dosing); significant tolerability issues.
• subjects initiate treatment within 2 business days.
• - Informed consent Medical history - includes: recording past and present illnesses and collection of the subjects demographic data (birth date, sex, and race).
• Adverse events are recorded starting from the time informed consent is obtained.
• Prior medications are collected from 4 weeks before the first dose of test article. All current medications are recorded.
• CBC Complete blood count
• WBC white blood cell count
• RBC red blood cell count
• Hb haemoglobin
• Hct hematocrit
• - Fasting chemistry panel/electrolytes includes sodium, potassium, chloride, blood urea nitrogen (BUN; or urea), serum creatinine, calculated clearance creatinine (CKD-EPI), glucose, calcium, phosphorus, total protein, uric acid, AST, ALT, ⁇ GT, ALP, total and direct bilirubine, albumin, total cholesterol, HDL, LDL, triglycerides, LDH, CPK, - ABG (for assessing respiratory and/or metabolic disorders)
• ApoA-I for pharmacokinetic and pharmacodynamic assessment
• Coagulation tests includes prothrombin time (PT) (expressed as international normalized ratio [INR]), and partial thromboplastin time (PTT).
• Urinalysis includes specific gravity, pH, assessment of protein/albumin, glucose, ketones, and haemoglobin/blood.
• Pharmacokinetic and pharmacodynamic assessment includes apoA-I and total cholesterol levels.
• Endotoxin levels are measured using the EAATM kit.
• AKI Biomarkers (TIMP-2 and IGFBP-7) are measured using the Nephrocheck® kit. Inflammatory markers include: CRP, D- dimer, Ferritin, IL-6, IL-8, GM-CSF, MCP 1 and TNF- ⁇ .
• CBC Complete blood count
• WBC white blood cell count
• RBC red blood cell count
• Hb haemoglobin
• Hct hematocrit
• - Fasting chemistry panel/electrolytes includes sodium, potassium, chloride, blood urea nitrogen (BUN; or urea), serum creatinine, calculated clearance creatinine (CKD-EPI), - glucose, calcium, phosphorus, total protein, uric acid, AST, ALT, ⁇ GT, ALP, total and direct bilirubine, albumin, total cholesterol, HDL, LDL, triglycerides, LDH, CPK - ABG (for assessing respiratory and/or metabolic disorders) - ApoA-I (for pharmacokinetic and pharmacodynamic assessment) - Coagulation tests – includes prothrombin time (PT) (expressed as international normalized ratio [INR]), and partial thromboplastin time (PTT).
• PT prothrombin time
• ITR international normalized ratio
• PTT partial thromboplastin time
• Urinalysis includes specific gravity, pH, assessment of protein/albumin, glucose, ketones, and haemoglobin/blood.
• Pharmacokinetic and pharmacodynamic assessment will include apoA-I and total cholesterol levels.
• Endotoxin levels are measured using the EAATM kit.
• AKI Biomarkers (TIMP-2 and IGFBP-7) are measured using the Nephrocheck® kit. Inflammatory markers include: CRP, D- dimer, Ferritin, IL-6, IL-8, GM-CSF, MCP 1 and TNF- ⁇ .
• Table 4 provides a summary of the study protocol of this Example.
• Safety evaluations are attained utilizing information collected from the following assessments: physical examination (including weight), vital signs (blood pressure, pulse, temperature), CBC with differential, platelet count, blood chemistries, and fasting lipid profiles [including HDL-cholesterol, LDL-cholesterol and Lipoprotein (a) ], urea, glucose, 24 hour urine protein determination, serum creatinine and calculated creatinine clearance (CKD- EPI) and adverse events monitoring. All women of childbearing potential have a qualitative serum pregnancy test during pre-study screening/baseline evaluation and subsequently, if clinically indicated. Patients are monitored throughout the study for the occurrence of adverse events, that are recorded.
• Adverse events volunteered by the subject or discovered, as a result of general questioning by the investigator or by physical examination, are recorded. The duration (start and end dates), severity, cause and relationship to study medication, patient outcome, action taken, and an assessment of whether the event was serious are recorded for each reported adverse event. [0226] Adverse events: Definitions [0227] The term “adverse event,” is synonymous with the term “adverse experience,” which is used by the FDA. An adverse event (AE) is any untoward, undesired, unplanned clinical event in the form of signs, symptoms, disease, or laboratory or physiological observations occurring in a human being participating in a clinical study regardless of causal relationship. This includes the following: - Any clinically significant worsening of a pre-existing condition.
• a procedure is not an AE, but the reason for a procedure may be an AE.
• a “preexisting condition” is a clinical condition (including a condition being treated) that is diagnosed before the subject signs the informed consent form and that is documented as part of the subject’s medical history.
• TEAE treatment-emergent adverse event
• a “serious adverse event” is any AE occurring at any dose that meets 1 or more of the following criteria: - Results in death - Is life-threatening (see below) - Requires in subject hospitalization or prolongation of an existing hospitalization (see below) - Results in a persistent or significant disability or incapacity (see below) - Results in a new malignancy - Results in a congenital anomaly or birth defect [0231] 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 may jeopardize the subject and may require medical or surgical intervention to prevent one of the outcomes listed above.
• a “life-threatening adverse event” is any AE that places the subject at immediate risk of death from the event as it occurred.
• a life-threatening event does not include an event that might have caused death had it occurred in a more severe form but that did not create an immediate risk of death as it actually occurred. For example, drug-induced hepatitis that resolved without evidence of hepatic failure would not be considered life-threatening, even though drug-induced hepatitis of a more severe nature can be fatal.
• Hospitalization or prolongation of a hospitalization is a criterion for considering an AE to be serious. In the absence of an AE, the participating investigator should not report hospitalization or prolongation of hospitalization on a form. This is the case in the following situations: Hospitalization or prolongation of hospitalization is needed for a procedure required by the protocol. Day or night survey visits required by the protocol are not considered serious. [0234] Timing for reporting serious adverse events: Any SAE, regardless of causal relationship, is reported to medical monitor immediately (no later than 24 hours after the investigator becomes aware of the SAE) by faxing a completed serious adverse event form.
• the subject is asked the following non specific question: “How have you been feeling since your last visit?” Signs and symptoms are recorded using standard medical terminology.
• the health outcomes assessment surveys administered to study subjects are intended to explore the subject’s own perceptions about their quality of life. However, the investigator reviews the survey for the presence of potential AEs or SAEs and considers the subject’s perceptions when determining the occurrence of an AE or SAE.
• the subject’s assessments are not intended to be influenced by the clinical investigator. Every effort is made to maintain an unbiased assessment.
• the following AE information is included (when applicable): the specific condition or event and direction of change; whether the condition was pre-existing (i.e.
• Event may be explained by administration of the test article, or by the subject’s clinical state or other agents/therapies.
• Event is most likely to be explained by the subject’s clinical state or other agents/therapies, rather than the test article.
• Event can be fully explained by the subject’s clinical state or other agents/therapies.
• a protocol-related SAE may be an event that occurs during a washout period or that is related to a procedure required by the protocol.
• the severity of AEs is assessed according to the National Cancer Institute (NCI) Common Toxicity Criteria for Adverse Events (CTCAE) version 5.0.
• NCI National Cancer Institute
• CTCAE Common Toxicity Criteria for Adverse Events
• the following definitions are used for toxicities that are not defined in the NCI CTCAE: - Mild (Grade 1): The AE is noticeable to the subject but does not interfere with routine activity. The AE does not require discontinuing administration or reducing the dose of the test article.
• - Moderate (Grade 2) The AE interferes with routine activity but responds to symptomatic therapy or rest. The AE may require reducing the dose but not discontinuing administration of the test article.
• COVID-19 is infects host cells through binding of the viral spike protein (SARS-2-S) to the cell-surface receptor angiotensin-converting enzyme 2 (ACE2), and the HDL scavenger receptor B type 1 (SR-B1) facilitates ACE2-dependent entry of the virus.
• SARS-2-S viral spike protein
• ACE2 cell-surface receptor angiotensin-converting enzyme 2
• SR-B1 HDL scavenger receptor B type 1
• lipid binding protein-based complexes such as CER-001 may provide a therapeutic benefit (e.g., reducing the severity and/or duration of CRS) in subjects having a COVID-19 infection through competitive binding to SR-B1, thereby limiting the virus’s ability to infect additional cells.
• CRS lipid binding protein-based complexes
• a pilot study is conducted to investigate the safety and efficacy of seven CER-001 infusions in patients with CRS secondary to COVID-19 infection. The study consists of 9 visits: • Pre-Dosing (Baseline) Visit: Assessment of baseline inflammatory markers and safety labs. • Dosing Visits: Seven doses (Doses 1 through 7) are administered as a once daily infusion over a 7-day period.
• IL-6 is measured daily from a pre-infusion sample. • follow-Up Visit: Patients have their final evaluation on Day 8. Inflammatory markers and safety labs are measured. [0251] A flowchart for the study is shown in FIG.6. 7.3.1. Selection of Study Subjects 7.3.1.1. Inclusion Criteria [0252] Eligible patients meeting the following criteria are enrolled into the study: 1. Male or non-pregnant female adult ⁇ 18 years of age at time of enrollment. 2. Has laboratory-confirmed novel coronavirus (COVID-19) infection as determined by polymerase chain reaction (PCR), or other commercial or public health assay in oropharyngeal or anal specimen within 72 hours prior to hospitalization. 3.
• COVID-19 novel coronavirus
• Illness of any duration and at least one of the following: a. Radiographic infiltrates by imaging (chest x-ray, CT scan, etc.), OR b. Clinical assessment (evidence of rales/crackles on physical examination) AND SpO2 ⁇ 93% on room air, OR c. Requiring mechanical ventilation and/or supplemental oxygen, OR d. Sustained fever in the past 24 hours and unresponsive to NSAID or steroid 4. Serum IL-6 ⁇ 3 times the upper limit of normal 5. Females of childbearing potential that agree and commit to use an acceptable form of birth control for the entire study.
• Pregnancy or breast feeding 8. Anticipated transfer to another hospital which is not a study site within 72 hours. 9. Expected life span does not exceed 7 days. 10. Patients who have used an investigational agent within 30 days of the first dose of CER-001. 7.3.1.2. Restrictions During the Study [0254] There are no patient restrictions other than those outlined in the Inclusion/Exclusion criteria above. 7.3.1.3.
• Reasons for withdrawal of a patient from study drug may include, but are not limited to, the following: • Investigator's request, for safety reasons, such as severe adverse reactions; • Investigator’s request, for other reasons, such as patient non-compliance; • Patient’s request, for tolerability reasons; • Patient’s request, for other reasons, such as withdrawal of informed consent; [0256] Discontinuation of study drug alone does not constitute discontinuation or withdrawal from the study. Patients continue to be followed as though they had completed the treatment phase. Patients who prematurely discontinue study medication (e.g., prior to completion of the 7th dose) undergo end of study evaluations whenever possible. 7.3.2. Treatment of Patients 7.3.2.1.
• CER-001 is provided 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 with normal saline to a volume of 250 mL. [0258] Dosing occurs at each of the seven dosing visits. At each of these visits, patients are given a single IV infusion CER 001 (20 mg/kg) over a period of 24 hours using an infusion pump. Patients are pretreated with antihistamine prior to each CER-001 dose (e.g. dexchlorpheniramine 5 mg or hydroxyzine 100 mg) to avoid any potential infusion reactions.
• antihistamine e.g. dexchlorpheniramine 5 mg or hydroxyzine 100 mg
• Efficacy Parameters (a) Primary Efficacy Parameters [0265] The primary efficacy parameter is the change in IL-6 from baseline to Day 8. Baseline is defined as the average of the measurements taken at the baseline visit and prior to dosing on Day 1. (b) Secondary Efficacy Parameters [0266] Secondary efficacy parameters include changes to the inflammatory markers CRP, D- dimer, Ferritin, IL-8, GM-CSF, MCP 1 and TNF- ⁇ from baseline to Day 8. 7.3.7. Assessment of Safety 7.3.7.1. Safety Parameters (a) Pregnancy Tests (if applicable) [0267] Females of child bearing potential have a documented negative pregnancy test performed any time during hospitalization and prior to dosing.
• Example 4 CER-001 Therapy for Treating CRS Secondary to Covid-19 Infection - Additional Treatment Protocol
• This Example is a study of CER-001 therapy in COVID-19 patients with severe cytokine release syndrome and renal injury. 7.4.1. Selection of Subjects 7.4.1.1. Inclusion Criteria [0271] Eligible patients meet the following criteria before they are enrolled into the study: 1. Male or non-pregnant female adult 18 years of age at time of enrollment. 2.
• PCR polymerase chain reaction
• Illness of any duration and at least one of the following: Radiographic infiltrates by imaging (chest x-ray, CT scan, etc.), OR Clinical assessment (evidence of rales/crackles on physical examination) AND SpO2 ⁇ 93% on room air, OR Requiring mechanical ventilation and/or supplemental oxygen, OR Sustained fever in the past 24 hours and unresponsive to NSAID or steroid 4. Serum IL-6 >3 times the upper limit of normal 5.
• Patients are pretreated with antihistamine prior to each CER-001 dose (e.g. dexchlorpheniramine 5 mg or hydroxyzine 100 mg) to avoid any potential infusion reactions.
• Patients receive IV infusion of CER-001 at the dosage of 15 mg/kg BID for 3 consecutive days. At the discretion of the investigator, patients may receive up to two additional doses.
• Patients may be interrupted or discontinued from study medication if any of the following occur: 1. Any drug-related adverse event or other reason which, in the Investigator's opinion, jeopardizes the patient's participation in the trial or the interpretation of trial data (e.g., severe inter-current illness requiring additional care measures or preventing further dosing); 2.
• Medical history includes recording past and present illnesses and collection of the subject’s demographic data (birth date, sex, and race). 3. Physical examination with a review of systems, height and weight, BMI and wait circumference 4. Vital signs (pulse, blood pressure, and oral, auricular, axillary, or core temperature). 5. Review of inclusion/exclusion criteria. 6. Adverse events are recorded starting from the time informed consent is obtained. 7. Prior medications are collected from 4 weeks before the first dose of test article. All current medications are recorded. 8. Complete blood count (CBC) – includes white blood cell count (WBC) with differential, platelet count, red blood cell count (RBC), haemoglobin (Hb), hematocrit (Hct). 9.
• WBC white blood cell count
• RBC red blood cell count
• Hb haemoglobin
• Hct hematocrit
• Fasting chemistry panel/electrolytes includes sodium, potassium, chloride, blood urea nitrogen (BUN; or urea), serum creatinine, calculated clearance creatinine (CKD-EPI), glucose, calcium, phosphorus, total protein, uric acid, AST, ALT, GT, ALP, total and direct bilirubine, albumin, total cholesterol, HDL, LDL, triglycerides, LDH, CPK, 10.
• ABG for assessing respiratory and/or metabolic disorders
• ApoA-I for pharmacokinetic and pharmacodynamic assessment
• Coagulation tests includes prothrombin time (PT) (expressed as international normalized ratio [INR]), and partial thromboplastin time (PTT). 13.
• Urinalysis includes specific gravity, pH, assessment of protein/albumin, glucose, ketones, and haemoglobin/blood. 14. Microalbumunuria and Proteinuria g/24 h 15. Serum or urine pregnancy test (for women of childbearing potential) within 7 days before randomization. 16. Pharmacokinetic and pharmacodynamic assessment include apoA-I and total cholesterol levels. 17. Inflammatory markers include CRP, PCT, D-dimer, Ferritin, IL-6, IL-8, GM-CSF, MCP 1 and TNF- ⁇ . [0282] Clinical and laboratory parameters are monitored from baseline to the Final visit at Day 8 as reported in FIG.7 and include the following procedures: 1. Recording of adverse events and concomitant medications 2. Review of appropriate laboratory information 3.
• CBC Complete blood count
• WBC white blood cell count
• RBC red blood cell count
• Hb haemoglobin
• Hct hematocrit
• Fasting chemistry panel/electrolytes includes sodium, potassium, chloride, blood urea nitrogen (BUN; or urea), serum creatinine, calculated clearance creatinine (CKD-EPI), glucose, calcium, phosphorus, total protein, uric acid, AST, ALT, GT, ALP, total and direct bilirubine, albumin, total cholesterol, HDL, LDL, triglycerides, LDH, CPK 8.
• ABG for assessing respiratory and/or metabolic disorders
• ApoA-I for pharmacokinetic and pharmacodynamic assessment
• Coagulation tests includes prothrombin time (PT) (expressed as international normalized ratio [INR]), and partial thromboplastin time (PTT). 11.
• Urinalysis includes specific gravity, pH, assessment of protein/albumin, glucose, ketones, and haemoglobin/blood. 12. Microalbumunuria and Proteinuria g/24 h 13. Inflammatory markers include CRP, PCT, D-dimer, Ferritin, IL-6, IL-8, GM-CSF, MCP 1 and TNF- ⁇ 7.4.4. Adverse Event (AE) Reporting [0283] An AE is any untoward medical occurrence associated with the use of the investigational product (active or placebo drug, biologic, or device) in a clinical investigation patient, which does not necessarily have a causal relationship with the product.
• Adverse events may include: • Symptoms described by the patient • Clinically significant changes in the patient’s physical exam or other signs observed by the Investigator or medical staff • Test abnormalities (laboratory tests) that reflect a change from baseline and/or that may result in changes in administration of investigational product or in an alteration in medical care (diagnostic or therapeutic) • Conditions present at baseline that have either worsened or recurred following resolution [0285] The patients are evaluated for new AEs and the status of existing AEs at each study visit. 7.4.5.
• Example 5 CER-001 Therapy for Treating Ischemia/reperfusion AKI
• This Example is a study of CER-001 therapy for treating ischemia/reperfusion AKI. 7.5.1. Materials and Methods [0288] Pigs, with a body weight of 45-60 kg, are fasted for 24 hours before the study. All animals are premedicated with an intramuscular mixture of azaperone (8 mg kg -1 ) and atropine (0.03 mg kg -1 ) to reduce pharyngeal and tracheal secretions and prevent post-intubation bradycardia. After anesthesia, both kidneys are approached through a midline abdominal incision.
• a method of treating a subject with an acute condition comprising administering to the subject in need thereof a high dose of a lipid binding protein-based complex, optionally wherein the acute condition comprises acute inflammation.
• the method of embodiment 1, wherein the high dose is administered over a period of three days to approximately two weeks, optionally wherein the high dose is administered over a period of three days, four days, five days, six days, seven days, eight days, nine days, 10 days, eleven days, 12 days, 13 days, 14 days or 15 days.
• the high dose is the aggregate of two to ten individual doses, optionally wherein the high dose is an aggregate of three, four, five, six, seven, eight, nine or 10 individual doses. 4.
• each individual dose is effective to increase the subject’s HDL levels by at least 25%, at least 30% or at least 35% 2-4 hours after administration. 8.
• each individual dose is effective to increase the subject’s HDL levels by at least 25%, at least 30% or at least 35% 2 hours after administration.
• each individual dose is effective to increase the subject’s HDL levels by at least 25%, at least 30% or at least 35% 3 hours after administration.
• each individual dose is effective to increase the subject’s HDL levels by at least 25%, at least 30% or at least 35% 4 hours after administration.
• 11. The method of any one of embodiments 3 to 10, wherein each individual dose is effective to increase the subject’s ApoA-I levels. 12.
• each individual dose is effective to increase the subject’s ApoA-I levels by at least 25%, at least 30% or at least 35% 2-4 hours after administration.
• each individual dose is effective to increase the subject’s ApoA-I levels by at least 25%, at least 30% or at least 35% 2 hours after administration.
• each individual dose is effective to increase the subject’s ApoA-I levels by at least 25%, at least 30% or at least 35% 3 hours after administration.
• each individual dose is effective to increase the subject’s ApoA-I levels by at least 25%, at least 30% or at least 35% 4 hours after administration.
• IL-6 interleukin-6
• the method of embodiment 17 or embodiment 18, wherein the high dose is effective to reduce serum levels of C-reactive protein.
• the high dose is effective to reduce serum levels of D-dimer. 21.
• TNF- ⁇ tumor necrosis factor ⁇
• 27. The method of any one of embodiments 17 to 26, wherein the high dose is effective to reduce serum levels of the one or more inflammatory markers by at least 20%, by at least 40% or by at least 60%.
• 28. The method of any one of embodiments 1 to 27, wherein the subject has CRS or is at risk of CRS. 29.
• 30 The method of embodiment 29, wherein the subject has CRS secondary to an infection.
• 31. The method of embodiment 30, wherein the infection is a viral infection.
• 32. The method of embodiment 31, wherein the viral infection is a coronavirus infection. 33.
• the method of embodiment 32, wherein the coronavirus is COVID-19.
• 34. The method of embodiment 31, wherein the viral infection is influenza infection.
• 35. The method of embodiment 29, wherein the subject has CRS caused by immunotherapy.
• 36. The method of embodiment 35, wherein the immunotherapy comprises antibody therapy.
• 37. The method of embodiment 35, wherein the immunotherapy comprises chimeric antigen receptor (CAR) T cell therapy.
• 38. The method of any one of embodiments 35 to 37, wherein the lipid binding protein-based complex is administered before the immunotherapy begins.
• 39. The method of any one of embodiments 35 to 38, wherein the lipid binding protein-based complex is administered concurrently with the immunotherapy. 40.
• the immunotherapy comprises chimeric antigen receptor (CAR) T cell therapy.
• CAR chimeric antigen receptor
• 53. The method of any one of embodiments 1 to 27, wherein the subject has or is at risk of developing sepsis.
• AKI acute kidney injury
• 65 The method of embodiment 62, wherein the AKI is cardiac surgery-associated AKI.
• the method of embodiment 62, wherein the AKI is hepatorenal syndrome (HRS) AKI.
• HRS hepatorenal syndrome
• the method of embodiment 66, wherein the HRS is type 2 HRS. 69.
• the method of embodiment 69, wherein the AKI is secondary to a viral infection, optionally wherein the viral infection is COVID-19.
• the method of embodiment 69 or embodiment 70, wherein the high dose is effective to reduce the severity of the AKI.
• the method of any one of embodiments 62 to 66, wherein the subject is at risk for AKI.
• the method of embodiment 73, wherein the sepsis is associated with a gram- negative bacterial infection. 75.
• the method of embodiment 72, wherein the subject has chronic liver disease. 82.
• the method of any one of embodiments 72 to 81, wherein the high dose is effective to reduce the likelihood that the subject will develop AKI. 83.
• the method of any one of embodiments 72 to 82, wherein the high dose is effective to delay the onset of AKI. 84.
• the method of any one of embodiments 72 to 82, wherein the high dose is effective to prevent AKI.
• lipid binding protein-based complex comprises a sphingomyelin.
• lipid binding protein-based complex comprises a negatively charged lipid.
• DPPG 1,2- dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol) (DPPG) or a salt thereof.
• DPPG 1,2- dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol) (DPPG) or a salt thereof.
• DPPG 1,2- dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol) (DPPG) or a salt thereof.
• lipid binding protein-based complex is CER-001. 101. The method of any one of embodiments 1 to 100, wherein the lipid binding protein-based complex is administered systemically, optionally by infusion. 102. The method of any one of embodiments 1 to 101, wherein the lipid binding protein-based complex is administered until serum levels of one or more inflammatory markers are reduced. 103. The method of embodiment 102, wherein the lipid binding protein-based complex is administered until serum levels of one or more inflammatory markers are reduced to a normal range(s). 104.
• lipid binding protein-based complex is administered until serum levels of one or more inflammatory markers are reduced below a baseline level(s) for the one or more inflammatory markers measured prior to lipid binding protein-based complex administration.
• 105. The method of any one of embodiments 1 to 104, wherein each individual dose of the lipid binding protein-based complex administered is 4-40 mg/kg (on a protein weight basis).
• 106. The method of embodiment 105, wherein each individual dose of the lipid binding protein-based complex is 4-30 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 15-25 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 10-30 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 10-20 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 5 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 10 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 15 mg/kg (on a protein weight basis). 113.
• each individual dose of the lipid binding protein-based complex is 20 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 5 to 15 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 10 to 20 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex is 15 to 25 mg/kg (on a protein weight basis).
• the induction regimen comprises administering the lipid binding protein-based complex once daily or twice daily.
• the consolidation regimen comprises administering the lipid binding protein-based complex once daily or once every two days.
• the subject is not treated with a maintenance regimen.
• the consolidation regimen comprises administering one or more doses of the lipid binding protein- based complex to the subject one or more days after administration of the final dose of the induction regimen. 122.
• the method of embodiment 121, wherein the first dose of the lipid binding protein-based complex administered during the consolidation regimen is administered two or more days after administration of the final dose of the induction regimen.
• the first dose of the lipid binding protein-based complex administered during the consolidation regimen is administered three or more days after administration of the final dose of the induction regimen.
• the first dose of the lipid binding protein-based complex administered during the consolidation regimen is administered three days after administration of the final dose of the induction regimen.
• any one of embodiments 117 to 124 which comprises an induction regimen comprising twice daily administration of the lipid binding protein-based complex on days 1, 2, and 3 and a consolidation regimen comprising two doses of the lipid binding protein-based complex on day 6.
• 126 The method of any one of embodiments 117 to 125, wherein each individual dose of the lipid binding protein-based complex administered in the induction regimen is 4-40 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered in the induction regimen is 4-30 mg/kg (on a protein weight basis). 128.
• each individual dose of the lipid binding protein-based complex administered in the induction regimen is 15-25 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered in the induction regimen is 10-30 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered in the induction regimen is 10-20 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered in the induction regimen is 5 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered in the induction regimen is 10 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered in the induction regimen is 15 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered in the induction regimen is 20 mg/kg (on a protein weight basis).
• 135. The method of any one of embodiments 117 to 134, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 5 to 15 mg/kg (on a protein weight basis).
• 136. The method of any one of embodiments 117 to 134, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 10 to 20 mg/kg (on a protein weight basis).
• the method of any one of embodiments 117 to 134, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 15 to 25 mg/kg (on a protein weight basis). 138. The method of any one of embodiments 117 to 134, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 5 mg/kg (on a protein weight basis). 139. The method of any one of embodiments 117 to 134, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 10 mg/kg (on a protein weight basis). 140. The method of any one of embodiments 117 to 134, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 15 mg/kg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 300 mg to 4000 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 300 mg to 3000 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 300 mg to 1500 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 400 mg to 4000 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 400 mg to 1500 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 500 mg to 1200 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 500 mg to 1000 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 600 mg to 3000 mg (on a protein weight basis). 149.
• each individual dose of the lipid binding protein-based complex administered is 800 mg to 3000 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 1000 mg to 2400 mg (on a protein weight basis).
• each individual dose of the lipid binding protein-based complex administered is 1000 mg to 2000 mg (on a protein weight basis).
• 152. The method of any one of embodiments 1 to 151, wherein the high dose of the lipid binding protein-based complex is 600 mg to 40 g (on a protein weight basis). 153.
• the method of any one of embodiments 1 to 151, wherein the high dose of the lipid binding protein-based complex is 3 g to 35 g (on a protein weight basis).
• 154. The method of any one of embodiments 1 to 151, wherein the high dose of the lipid binding protein-based complex is 5 g to 30 g (on a protein weight basis).
• 155. The method of any one of embodiments 1 to 154, wherein the lipid binding protein-based complex is administered by infusion.
• 156. The method of embodiment 155, wherein each individual dose is administered over a one to 24-hour period.
• the method of any one of embodiments 1 to 157 which further comprises administering an antihistamine to the subject prior to each individual dose.
• the antihistamine comprises dexchlorpheniramine or hydroxyzine.
• the subject is receiving or has received one or more additional therapies and/or which further comprises administering to the subject one or more additional therapies.
• the one or more additional therapies comprises one or more anti-IL-6 agents. 162.
• 164. The method of any one of embodiments 160 to 163, wherein the one or more additional therapies comprise one or more corticosteroids.
• the one or more corticosteroids comprise methylprednisolone, dexamethasone, or a combination thereof.
• the method of any one of embodiments 160 to 165, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise antibodies from recovered COVID-19 patients.
• the method of any one of embodiments 160to 166, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise antibodies against the spike protein of COVID-19.
• the method of any one of embodiments 160 to 167, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise one or more antiviral agents.
• the method of embodiment 168, wherein the one or more antiviral agents comprise lopinavir. 170.
• the method of embodiment 168 or embodiment 169, wherein the one or more antiviral agents comprise remdesivir. 171.
• the method of any one of embodiments 168 to 170, wherein the one or more antiviral agents comprise danoprevir. 172.
• the method of any one of embodiments 168 to 171, wherein the one or more antiviral agents comprise galidesivir. 173.
• the method of any one of embodiments 168 to 172, wherein the one or more antiviral agents comprise darunavir. 174.
• the method of any one of embodiments 168 to 173, wherein the one or more antiviral agents comprise ritonavir. 175.
• the method of any one of embodiments 160 to 174, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise chloroquine or hydroxychloroquine. 176.
• the method of any one of embodiments 160 to 175, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise azithromycin. 177.
• the method of any one of embodiments 160 to 176, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise an interferon.
• the method of embodiment 177, wherein the interferon is an interferon alpha. 179.
• the method of embodiment 177, wherein the interferon is an interferon beta. 180.
• the CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids in a ApoA-I weight:total phospholipid weight ratio of 1:2.7 +/- 20% and the phospholipids sphingomyelin and DPPG in a sphingomyelin:DPPG weight:weight ratio of 97:
• the CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids in a ApoA-I weight:total phospholipid weight ratio of 1:2.7 +/- 10% and the phospholipids sphingomyelin and DPPG in a sphingomyelin:DPPG weight:weight ratio of 97:3 +/- 10%.
• the CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids in a ApoA-I weight:total phospholipid weight ratio of 1:2.7 +/- 10% and the phospholipids sphingomyelin and DPPG in a sphingomyelin:DPPG weight:weight ratio of 97:3 +/- 10%.
• the method of any one of embodiments 182 to 186, wherein the CER-001 comprises natural sphingomyelin. 188.
• the method of embodiment 187, wherein the natural sphingomyelin is chicken egg sphingomyelin.
• the method of any one of embodiments 182 to 186, wherein the CER-001 comprises synthetic sphingomyelin.
• the method of embodiment 189, wherein the synthetic sphingomyelin is palmitoylsphingomyelin. 191.
• the method of any one of embodiments 181 to 190, wherein CER-001 is administered in the form of a formulation in which the CER-001 is at least 95% homogeneous. 192.
• a method of treating a subject with cytokine release syndrome (CRS) or at risk of CRS comprising administering to the subject a therapeutically effective amount of CER-001.
• the method of embodiment 1 which comprises administering an amount of CER-001 effective to reduce serum levels of one or more inflammatory markers in the subject.
• a method of reducing serum levels of one or more inflammatory markers in a subject in need thereof comprising administering to the subject an amount of CER-001 effective to reduce the serum levels of the one or more inflammatory markers.
• the method of any one of embodiments 1 to 4 wherein the subject has CRS. 6.
• the method of embodiment 6, wherein the infection is a viral infection.
• the viral infection is a coronavirus infection.
• the coronavirus is COVID-19.
• the method of embodiment 7, wherein the viral infection is influenza infection.
• the method of embodiment 5, wherein the subject has CRS caused by immunotherapy.
• the method of embodiment 11, wherein the immunotherapy comprises antibody therapy.
• the method of embodiment 11, wherein the immunotherapy comprises chimeric antigen receptor (CAR) T cell therapy.
• CAR chimeric antigen receptor
• CAR chimeric antigen receptor
• 26. The method of any one of embodiments 23 to 25, wherein CER-001 is administered before the immunotherapy begins.
• 27. The method of any one of embodiments 23 to 26, wherein CER-001 is administered concurrently with the immunotherapy.
• 28. The method of any one of embodiments 23 to 27, wherein CER-001 is administered after the immunotherapy ends.
• 29. The method of any one of embodiments 1 to 28, which comprises once daily administration of CER-001.
• 30. The method of any one of embodiments 1 to 29, wherein the CER-001 is administered for at least 5 days.
• 31. The method of any one of embodiments 1 to 29, wherein the CER-001 is administered for at least 6 days. 32.
• the method of embodiment 39, wherein the dose of CER-001 administered in the induction regimen is 20 mg/kg (on a protein weight basis). 43.
• the method of any one of embodiments 1 to 42, wherein the dose of CER-001 administered at each administration is 600 mg to 4000 mg. 44.
• the method of embodiment 43, wherein the dose of CER-001 administered at each administration is 600 mg to 3000 mg. 45.
• the method of embodiment 43, wherein the dose of CER-001 administered at each administration is 800 mg to 3000 mg. 46.
• the method of embodiment 43, wherein the dose of CER-001 at each administration is 1000 mg to 2400 mg. 47.
• the method of embodiment 43, wherein the dose of CER-001 administered at each administration is 1000 mg to 2000 mg. 48.
• the method of any one of embodiments 2 to 50, wherein the one or more inflammatory markers comprise interleukin 6 (IL-6).
• the method of any one of embodiments 2 to 51, wherein the one or more inflammatory markers comprise C-reactive protein.
• the method of any one of embodiments 2 to 52, wherein the one or more inflammatory markers comprise D-dimer.
• 54. The method of any one of embodiments 2 to 53, wherein the one or more inflammatory markers comprise ferritin. 55.
• IL-8 interleukin 8
• GM-CSF granulocyte-macrophage colony stimulating factor
• MCP monocyte chemoattractant protein
• the antihistamine comprises dexchlorpheniramine or hydroxyzine.
• 61. The method of any one of embodiments 1 to 60, wherein the subject is receiving or has received one or more additional therapies and/or which further comprises administering to the subject one or more additional therapies.
• 62. The method of embodiment 61, wherein the one or more additional therapies comprises one or more anti-IL-6 agents.
• 63. The method of embodiment 62, wherein the one or more anti-IL-6 agents comprise tocilizumab, siltuximab, olokizumab, elsilimomab, BMS-945429, sirukumab, levilimab, CPSI-2364, or a combination thereof.
• the method of any one of embodiments 61 to 67, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise antibodies against the spike protein of COVID-19.
• the method of any one of embodiments 61 to 68, wherein the subject has or has had a COVID-19 infection and the one or more additional therapies comprise one or more antiviral agents.
• the one or more antiviral agents comprise lopinavir. 71.
• the method of embodiment 69 or embodiment 70, wherein the one or more antiviral agents comprise remdesivir.
• 72 The method of any one of embodiments 69 to 71, wherein the one or more antiviral agents comprise danoprevir. 73.
• a method of treating a subject with sepsis comprising administering to the subject an amount of a lipid binding protein-based complex.
• the method of embodiment 1 or embodiment 2 wherein the subject has an intra-abdominal cavity infection.
• the method of any one of embodiments 1 to 4 wherein the amount of the lipid binding protein-based complex is effective to reduce the severity of the sepsis.
• the method of any one of embodiments 1 to 5 wherein the amount of the lipid binding protein-based complex is effective to reduce the likelihood that the subject will develop acute kidney injury (AKI). 7.
• AKI acute kidney injury
• a method of treating a subject with acute kidney injury (AKI) or at risk for AKI comprising administering to the subject an amount of a lipid binding protein-based complex. 10. The method of embodiment 9, wherein the AKI is sepsis-related AKI. 11. The method of embodiment 9 or embodiment 10, wherein the subject has AKI. 12. The method of embodiment 11, wherein the amount of the lipid binding protein- complex is effective to reduce the severity of the AKI. 13.
• a method of treating a subject with acute kidney injury (AKI) or at risk for AKI comprising administering to the subject an amount of a lipid binding protein-based complex.
• AKI is sepsis-related AKI.
• the method of embodiment 9 or embodiment 10 wherein the subject has AKI.
• the amount of the lipid binding protein- complex is effective to reduce the severity of the AKI. 13.
• the method of embodiment 9 or embodiment 10, wherein the subject is at risk for AKI.
• 14. The method of embodiment 13, wherein the subject has sepsis. 15.
• the method of embodiment 14, wherein the sepsis is associated with a gram- negative bacterial infection.
• 16. The method of embodiment 14 or embodiment 15, wherein the subject has an intra-abdominal cavity infection.
• 17. The method of embodiment 14 or embodiment 15, wherein the subject has urosepsis.
• 18. The method of any one of embodiments 13 to 17, wherein the amount of the lipid binding protein-based complex is effective to reduce the likelihood that the subject will develop AKI.
• 19. The method of any one of embodiments 13 to 18, wherein the amount of the lipid binding protein-based complex is effective to delay the onset of AKI. 20.
• 33. The method of any one of embodiments 1 to 32, wherein the lipid binding protein-based complex comprises a sphingomyelin.
• the negatively charged lipid is 1,2- dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol) (DPPG) or a salt thereof.
• DPPG 1,2- dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol) (DPPG) or a salt thereof.
• the lipid binding protein-based complex is CER-001, CSL-111, CSL-112, CER-522 or ETC-216.
• the lipid binding protein-based complex is CER-001.
• 38. The method of any one of embodiments 1 to 37, wherein the lipid binding protein-based complex is administered systemically, optionally by infusion. 39.
• lipid binding protein-based complex is administered according to a dosing regimen which comprises: (a) an induction regimen; and, optionally (b) a consolidation regimen, optionally wherein the lipid binding protein-based complex comprises CER-001.
• a dosing regimen which comprises: (a) an induction regimen; and, optionally (b) a consolidation regimen, optionally wherein the lipid binding protein-based complex comprises CER-001.
• the induction regimen comprises administering the lipid binding protein-based complex on multiple consecutive days.
• the induction regimen comprises administering the lipid binding protein-based complex on three or more consecutive days.
• 42. The method of any one of embodiments 39 to 41, wherein the induction regimen comprises twice daily administration of the lipid binding protein-based complex. 43.
• the induction regimen comprises twice daily administration of the lipid binding protein-based complex for three consecutive days.
• the dose of the lipid binding protein-based complex administered in the induction regimen is 4 to 30 mg/kg (on a protein weight basis).
• the dose of the lipid binding protein-based complex administered in the induction regimen is 5 to 15 mg/kg (on a protein weight basis).
• the dose of the lipid binding protein-based complex administered in the induction regimen is 10 to 20 mg/kg (on a protein weight basis). 47.
• the method of any one of embodiments 39 to 43, wherein the dose of the lipid binding protein-based complex administered in the induction regimen is 15 to 25 mg/kg (on a protein weight basis).
• 48. The method of any one of embodiments 39 to 43, wherein the dose of the lipid binding protein-based complex administered in the induction regimen is 5mg/kg (on a protein weight basis).
• 50. The method of any one of embodiments 39 to 43, wherein the dose of the lipid binding protein-based complex administered in the induction regimen is 20 mg/kg (on a protein weight basis). 51.
• the method of any one of embodiments 39 to 43, wherein the dose of CER-001 administered in the induction regimen is 300 mg to 3000 mg. 52.
• the method of any one of embodiments 39 to 43, wherein the dose of CER-001 administered in the induction regimen is 300 mg to 1500 mg. 53.
• the method of any one of embodiments 39 to 43, wherein the dose of CER-001 administered in the induction regimen is 400 mg to 1500 mg. 54.
• the method of any one of embodiments 39 to 43, wherein the dose of CER-001 administered in the induction regimen is 500 mg to 1200 mg. 55.
• the method of any one of embodiments 39 to 43, wherein the dose of CER-001 administered in the induction regimen is 500 mg to 1000 mg. 56.
• any one of embodiments 39 to 55 which comprises a consolidation regimen.
• the consolidation regimen comprises administering one or more doses of the lipid binding protein-based complex to the subject one or more days after administration of the final dose of the induction regimen.
• the first dose of the lipid binding protein- based complex administered during the consolidation regimen is administered two or more days after administration of the final dose of the induction regimen.
• the first dose of the lipid binding protein- based complex administered during the consolidation regimen is administered three or more days after administration of the final dose of the induction regimen. 60.
• the method of any one of embodiments 39 to 62, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 4 to 30 mg/kg (on a protein weight basis).
• the method of any one of embodiments 39 to 62, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 5 to 15 mg/kg (on a protein weight basis).
• the method of any one of embodiments 39 to 62, wherein the dose of the lipid binding protein-based complex administered in the consolidation regimen is 10 to 20 mg/kg (on a protein weight basis).
• the dose of the lipid binding protein-based complex administered in the consolidation regimen is 15 to 25 mg/kg (on a protein weight basis).
• the method of any one of embodiments 39 to 62, wherein the dose of CER-001 administered in the induction regimen is 300 mg to 1500 mg. 72.
• the method of any one of embodiments 39 to 62, wherein the dose of CER-001 administered in the induction regimen is 400 mg to 1500 mg. 73.
• the method of any one of embodiments v, wherein the dose of CER-001 administered in the induction regimen is 500 mg to 1200 mg. 74.
• the method of any one of embodiments 39 to 62, wherein the dose of CER-001 administered in the induction regimen is 500 mg to 1000 mg. 75.
• any one of embodiments 56 to 74 wherein the dose of the lipid binding protein-based complex administered in the induction regimen and the dose of the lipid binding protein-based complex administered in the consolidation regimen are the same.
• 76. The method of any one of embodiments 1 to 75, wherein an antihistamine is administered prior to administration of one or more of the lipid binding protein-based complex doses.
• 77. The method of embodiment 76, wherein an antihistamine is administered prior to each lipid binding protein-based complex dose.
• 78 The method of any one of embodiments 1 to 77, wherein the subject is also treated with a standard of care therapy for sepsis. 79.

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