Dosage regimens and related compositions and methods

Abstract

The present invention provides compounds and compositions for inhibiting complement-mediated damage to cells, tissues, or organs. [Solution] The present invention provides cell-reactive compstatin analogs and compositions comprising cell-reactive compstatin analogs. Furthermore, it provides methods for using cell-reactive compstatin analogs, for example, to inhibit complement-mediated damage to cells, tissues, or organs, for example, to treat complement-mediated disorders. Furthermore, it provides long-acting compstatin analogs and compositions comprising long-acting compstatin analogs. Furthermore, it provides methods for using long-acting compstatin analogs, for example, to inhibit complement-mediated damage to cells, tissues, or organs, for example, to treat complement-mediated disorders. Furthermore, it provides targeted compstatin analogs and compositions comprising targeted compstatin analogs.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims priority to U.S. Provisional Patent Application 62 / 599,637 filed December 15, 2017, U.S. Provisional Patent Application 62 / 655,094 filed April 9, 2018, and U.S. Provisional Patent Application 62 / 744,565 filed October 11, 2018, and incorporates the entire contents of each of these applications herein by reference. [Background technology]

[0002] background The complement system is a system of over 30 plasma and cell-binding proteins that plays a significant role in both innate and adaptive immunity. Complement proteins act in a series of enzymatic cascades involving diverse protein interactions and cleavage events. Complement activation occurs via three main pathways: the antibody-dependent classical pathway, the alternative pathway, and the mannose-binding lectin (MBL) pathway. Inappropriate or excessive complement activation is an underlying cause or factor in many serious diseases and conditions, and considerable effort has been made over the past few decades to explore various complement inhibitors as therapeutic agents. [Overview of the Initiative]

[0003] Summary of the Invention In some aspects, the present invention provides and / or relates to certain long-acting compstatin analogs. For example, the present invention provides and / or relates to such long-acting compstatin analogs, compositions comprising them, and methods for producing, identifying, characterizing, and / or using them. In some aspects, the present invention provides and / or relates to physiologically acceptable compositions comprising long-acting compstatin analogs. In some aspects, the present invention provides and / or relates to pharmaceutical-grade compositions comprising long-acting compstatin analogs. In particular, in some aspects, the present disclosure describes particularly useful long-acting compstatin analogs and further provides specific doses, forms of administration, administration regimens, unit dose compositions, and other technologies relating to the administration of long-acting compstatin analogs to human subjects, e.g., specific human subjects having and / or being susceptible to one or more certain diseases, disorders or conditions.

[0004] In one embodiment, the present invention provides a method for treating complement-mediated disorders in a subject requiring treatment, which may include administering a long-acting compstatin analog to the subject using a specific dose, a form of administration (e.g., a unit dose composition and / or a specific formulation) and / or an administration regime (determined as desired for the treatment of a particular disease, disorder, or condition, and in one embodiment, for example, a route of administration, timing of administration, etc.).

[0005] In some embodiments, the complement-mediated disorder treated according to the present invention is paroxysmal nocturnal hemoglobinuria (PNH), atypical hemolytic uremic syndrome (aHUS), or other disorders associated with complement-mediated hemolysis. In some embodiments, the disorder is ischemia / reperfusion (I / R) injury (e.g., myocardial infarction, thromboembolic stroke, or surgery-related). In some embodiments, the disorder is trauma. In some embodiments, the disorder is graft rejection. In some embodiments, the disorder is a chronic respiratory disorder, e.g., asthma or COPD.

[0006] All references, books, patent applications, patents, other publications, websites, and databases mentioned herein are incorporated herein by reference. In the event of any conflict between the statements herein and the incorporated materials, the statements herein (including any amendments thereof) shall prevail. Unless otherwise specified, the meanings of terms and abbreviations recognized in the art are used herein. The implementations described herein may employ conventional techniques that are within the ordinary art of molecular biology, cell culture, recombinant nucleic acid (e.g., DNA) techniques, immunology, and / or nucleic acid and polypeptide synthesis, detection, manipulation, and quantification. For example, Ausubel, F., et al., (eds.), Current Protocols in Molecular Biology, Current Protocols in Immunology, Current Protocols in Protein Science, and Current Protocols in Cell Biology, all John Wiley & Sons, NY, e.g., the current edition or subsequent editions as of January 2010; Sambrook, Russell, and Sambrook, Molecular Cloning: A Laboratory Manual, 3 rd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, 2001 or 4 th See ed., 2012. [Brief explanation of the drawing]

[0007] [Figure 1]This plot shows the percentage inhibition rate of complement activation by the compstatin analog CA28 (SEQ ID NO: 28) and three long-acting compstatin analogs (CA28-1, CA28-2, and CA28-3) as a function of peptide concentration (μM). Inhibition of complement activation was tested in vitro using a classical complement inhibition assay. The plot shows values ​​obtained by averaging the results of two measurements. CA28 (circles; red), CA28-1 (crosses (×); blue); CA28-2 (triangles; green), CA28-3 (squares; purple).

[0008] [Figure 2] This plot shows the percentage inhibition rate of complement activation by CA28 and its long-acting compstatin analogs CA28-2 and CA28-3 as a function of compound concentration (μM). CA28 (square, light gray), CA28-2 (diamond, black), CA28-3 (circle, dark gray). CA28-3 is a compound containing multiple peptide moieties. The activity per peptide moiety is lower than the activity of individual CA28 molecules, but the total activity of CA28-3 exceeds the activity of CA28 on a molar basis.

[0009] [Figure 3] This plot shows the plasma concentrations versus time of CA28 and the long-acting compstatin analogs CA28-2 and CA28-3 after a single intravenous injection in cynomolgus monkeys. CA28 was administered at 200 mg / kg. CA28-2 and CA28-3 were administered at 50 mg / kg each. In calculating these experimental doses, it was estimated that the administered CA28-2 and CA28-3 substances consisted of 80 w / w% active compound based on dry weight. However, during sample analysis, the standard curve was estimated to be 100 w / w% compound based on dry weight and estimated to be 30%. Therefore, the Cmax value is overestimated compared to the actual Cmax. CA28 (square, light gray), CA28-2 (triangle, black), CA28-3 (circle, dark gray).

[0010] [Figure 4]This plot shows the complement activation inhibition rate (percentage) of CA28 and the long-acting compstatin analog CA28-4 as a function of compound concentration (μM). Inhibition of complement activation was tested in vitro using a classical complement inhibition assay. The plot shows values ​​obtained by averaging the results of four measurements of CA28-4. CA28 (squares, light gray), CA28-4 (cross marks, black).

[0011] [Figure 5] This plot shows the plasma concentrations versus time of CA28 and long-acting compstatin analogs CA28-2, CA28-3, and CA28-4 after a single intravenous injection in cynomolgus monkeys. CA28 was administered at 200 mg / kg. CA28-2, CA28-3, and CA28-4 were each administered at 50 mg / kg. In calculating the dosages for these experiments, it was estimated that the administered CA28-2 and CA28-3 substances consisted of 80 w / w% active compounds based on dry weight. However, during sample analysis, the standard curve was estimated to be 100 w / w% of the compound based on dry weight. Therefore, the Cmax values ​​are estimated to be approximately 30% higher than the Cmax achieved when these compounds are administered at the doses shown on a dry weight basis. CA28 (square, light gray), CA28-2 (triangle, black), CA28-3 (circle, dark gray), CA28-4 (inverted triangle, black).

[0012] [Figure 6] This is a representative chromatogram showing the ultraviolet (UV) detection of a PEG-based long-acting compstatin analog using reversed-phase HPLC. The peak with a retention time (RT) of 33.68 minutes represents the pegylated compstatin analog and has a relative area of ​​96%.

[0013] [Figure 7]This plot shows the complement activation inhibitory activity percentage of CA28 and its long-acting compstatin analogs, CA28-2CS, CA28-2GS, CA28-2HS, and CA28-2TS, as a function of compound concentration (μM). CA28-2CS (diamond, red); CA28-2GS (cross, blue); CA28-2HS (triangle, green); CA28-2TS (square, black).

[0014] [Figure 8] This plot shows the complement activation inhibitory activity percentage of CA28 and its bifunctional long-acting compstatin analog, CA28-2GS-BF, as a function of compound concentration (μM). CA28 (hollow circles, blue); CA28-2G-SBF (filled circles, red).

[0015] [Figure 9] This plot shows the plasma concentrations versus time of CA28 and the long-acting compstatin analog CA28-2GS-BF in cynomolgus monkeys after administration by single intravenous injection (CA28 (square, red) and CA28-2GS-BF (circle, purple)) or once daily subcutaneous injection for 7 days (CA28-2GS-BF only, asterisk, blue). CA28-2GS-BF was administered at 25 mg / ml. The administration volume was 2 ml / kg intravenously and 0.28 ml / kg / day subcutaneously. The CA28 data were from different experiments, where the compound was also in 5% dextrose and formulated at 20 mg / ml in a 10 ml / kg administration volume. The medium for each case was an aqueous solution of 5% dextrose.

[0016] [Figure 10(A)] This plot shows the percentage of complement activation inhibitory activity of CA28 and the bifunctional long-acting compstatin analog, CA28-2TS-BF, as a function of compound concentration (μM). (A) Classical pathway inhibited by CA28 (circles, red) and CA28-2TS-BF (cross marks, blue).

[0017] [Figure 10(B)]This plot shows the complement activation inhibitory activity percentage of CA28 and its bifunctional long-acting compstatin analog, CA28-2TS-BF, as a function of compound concentration (μM). (B) Alternative pathway inhibition. CA28 (circles, red) and CA28-2TS-BF (cross marks, blue).

[0018] [Figure 10(C)] The structure of CA28-2TS-BF is shown (assuming the PEG portion is 40kD).

[0019] [Figure 11] This plot shows the plasma concentrations versus time of CA28 and the long-acting compstatin analog CA28-2TS-BF in cynomolgus monkeys after a single intravenous injection of CA28 at 200 mg / kg (square, red), a single intravenous injection of CA28-2TS-BF at 7 mg / kg (asterisk, purple), a single subcutaneous injection of CA28-2TS-BF at 7 mg / kg (circle, blue), or subcutaneous injection of CA28-2TS-BF at 7 mg / kg once daily for 7 consecutive days (inverted triangle, green). The medium for each case is an aqueous solution of 5% dextrose.

[0020] [Figure 12(A)] This shows flow cytometry analysis of C3 deposition on erythrocytes from PNH patients exposed to activated complement in a modified Ham test. It also shows the results of dilution experiments demonstrating the effect of CA28 on C3 deposition.

[0021] [Figure 12(B)] Flow cytometry analysis of C3 deposition on erythrocytes from PNH patients exposed to activated complement in a modified Ham test is shown. Dilution experiments demonstrating the effect of CA28-2GS-BF on C3 deposition are also shown. The concentrations of the compounds used are indicated in and above each panel.

[0022] [Figure 13]Flow cytometry analysis of C3 deposition on erythrocytes from patients with PNH who were exposed to activated complement in a modified Ham test in the absence of complement inhibitors (left panel), in the presence of the anti-C5 monoclonal antibody eculizumab (center panel), and in the presence of CA28-2GS-BF (right panel).

[0023] [Figure 14] This plot shows ex vivo serum-induced hemolysis observed in multiple dose-escalation clinical trials of long-acting compstatin analogs containing 40 kD PEG in healthy subjects.

[0024] [Figure 15] A schematic diagram of the target-mediated pharmacokinetic (TMDD) model is shown.

[0025] [Figure 16] The observed and predicted trough LACA-40 serum concentrations over baseline time, separated by administration regimen, are shown for both single-dose (SD) and 28-day multi-dose (MD, once daily) studies in healthy subjects. Black circles represent the mean PK concentration at each baseline time point, covered by the TMDD model prediction, represented by a solid line. Each administration regimen is shown in a different color. LACA-40 was administered subcutaneously.

[0026] [Figure 17] Predicted trough PK serum concentrations over baseline time after multiple once-daily (Q1D), subcutaneous (SC) administrations of LACA-40 at three dose levels: 180 mg, 270 mg, and 360 mg Q1D. Model predictions were based on the TMDD model.

[0027] [Figure 18] This chart shows predicted trough PK serum concentrations over baseline time after multiple once-daily (Q1D) and three-times-weekly (Monday / Wednesday / Friday) SC administrations of various doses of LACA-40. Each administration regimen is represented by a different colored line, and the model predictions are based on the TMDD model.

[0028] [Figure 19] This chart shows predicted trough PK serum concentrations after multiple once-daily (Q1D) and twice-weekly (Monday / Thursday) SC administrations of various doses of LACA-40. Each administration regimen is represented by a different colored line, and the model predictions are based on the TMDD model.

[0029] [Figure 20] This chart shows predicted trough PK serum concentrations over baseline time after multiple once-daily (Q1D) and once-weekly (Q1W) SC administrations of various doses of LACA-40. Each administration regimen is represented by a different colored line, and the model predictions are based on the TMDD model.

[0030] [Figure 21] This chart shows predicted trough PK serum concentrations over baseline time after multiple once-daily (Q1D) and once-weekly (Q1W) SC administrations of various doses of LACA-40. Each administration regimen is represented by a different colored line, and the model predictions are based on the TMDD model.

[0031] [Figure 22] The predicted serum concentrations over a 28-day administration period are shown after multiple once-daily (qd), twice-weekly (biw), or once-weekly (qw) SC administrations of various doses of LACA-40 corresponding to the administration regimens for cohorts 1, 2, and 3 in the clinical trials described in Example 30. Each administration regimen is represented by a different colored line, and the model predictions are based on the TMDD model. For the twice-weekly regimens, the plots reflect administrations at alternating 3-day and 4-day intervals (i.e., administrations on days 1, 4, 8, 11, 15, 18, 22, and 25).

[0032] [Figure 23] This shows extended forecasts up to 35 days for LACA-40 concentrations based on daily and twice-weekly administration during the 28-day period.

[0033] [Figure 24]The individual and mean summary observed serum concentrations of LACA-40 for cohorts 1 and 2 in the clinical trial described in Example 30 are shown.

[0034] [Figure 25] The individual and mean summary observed serum concentrations of LACA-40 for Cohort 1 in the clinical trial described in Example 30, as well as the predicted serum concentrations for this dosing regimen based on the TMDD model, are shown.

[0035] [Figure 26] Individual and mean summary observed LACA-40 serum concentrations for Cohort 2 in the clinical trial described in Example 30, as well as predicted serum concentrations for this dosing regimen based on the TMDD model, are shown.

[0036] [Figure 27] The mean summary observed LACA-40 serum concentrations for cohorts 1 and 2 in the clinical trial described in Example 30, and the predicted serum concentrations for these dosing regimens based on the TMDD model are shown. The plots further show that the observed PK data are consistent with the predictions from the TMDD model.

[0037] [Figure 28] The mean summary observed LACA-40 serum concentrations for cohorts 1 and 2 in the clinical trial described in Example 30, as well as the serum concentration for cohort 3 based on the TMDD model, are shown.

[0038] [Figure 29]The predicted serum concentrations over a 28-day administration period are shown after multiple once-daily (qd), twice-weekly (biw), or once-weekly (qw) SC administrations of various doses of LACA-40 corresponding to the administration regimens for cohorts 1, 2, 3, and 4 in the clinical trials described in Example 30. Each administration regimen is represented by a different colored line, and the model predictions are based on the TMDD model. For the twice-weekly regimens, the plots reflect administrations at alternating 3-day and 4-day intervals (i.e., administrations on days 1, 4, 8, 11, 15, 18, 22, and 25).

[0039] [Figure 30] The individual and mean summary observed serum concentrations of LACA-40 for cohorts 1, 2, 3, and 4 in the clinical trial described in Example 30 are shown.

[0040] [Figure 31] The individual and mean summary observed LACA-40 serum concentrations for Cohort 3 in the clinical trial described in Example 30, and the predicted serum concentrations for this dosing regimen based on the TMDD model are shown. The plot further shows that the observed PK data are consistent with the predictions from the TMDD model.

[0041] [Figure 32] The individual and mean summary observed LACA-40 serum concentrations for Cohort 4 in the clinical trial described in Example 30, and the predicted serum concentrations for this dosing regimen based on the TMDD model are shown. The plot further shows that the observed PK data are consistent with the predictions from the TMDD model. [Modes for carrying out the invention]

[0042] Detailed Description of an Embodiment of the Invention I. Definition The terms “approximately” or “about” in relation to numbers generally include numbers that fall within ±10%, in one embodiment ±5%, in another embodiment ±1%, or in another embodiment ±0.5% of the number, unless otherwise specified or otherwise evident from the context (unless such numbers exceed 100% of the possible number to an unacceptable degree).

[0043] Complement components or complement proteins (CRPs) are proteins involved in the activation of the complement system or in one or more complement-mediated activities. Components of the classical complement pathway include, for example, the C1q, C1r, C1s, C2, C3, C4, C5, C6, C7, C8, C9, and C5b-9 complexes (also known as membrane invasion complexes (MACs)) and any of the active fragments or enzymatic cleavage products of the aforementioned (e.g., C3a, C3b, C4a, C4b, C5a, etc.). Components of alternative pathways include, for example, factor B, factor D, and propagin. Components of the lectin pathway include, for example, MBL2, MASP-1, and MASP-2. Complement components also include cell-binding receptors for soluble complement components, such receptors mediating one or more of the biological activities of such soluble complement components after their binding. Such receptors include, for example, the C5a receptor (C5aR), the C3a receptor (C3aR), complement receptor 1 (CR1), complement receptor 2 (CR2), and complement receptor 3 (CR3, also known as CD45). It should be recognized that the term “complement components” is not intended to include molecules and molecular structures that act as “triggers” for complement activation, such as antigen-antibody complexes, or foreign structures found on microorganisms or artificial surfaces.

[0044] "Complement-mediated disorders" are any disorders in which complement activation is known or suspected to be a contributing factor and / or at least partially a causative factor in at least some subjects having such disorders, for example, disorders in which complement activation leads to tissue damage. Non-limiting examples of complement-mediated disorders include: (i) various disorders characterized by hemolysis or hemolytic anemia, such as atypical hemolytic uremic syndrome, warm antibody autoimmune hemolytic anemia, cold agglutinin disease, paroxysmal nocturnal hemoglobinuria, and transfusion reactions; (ii) graft rejection (e.g., hyperacute or acute graft rejection) or graft dysfunction; (iii) disorders including ischemia / reperfusion injury, such as trauma, surgery (e.g., aneurysm repair), myocardial infarction, and ischemic stroke; (iv) respiratory disorders, such as asthma and chronic obstructive pulmonary disease (COPD); (v) arthritis, such as rheumatoid arthritis; and (vi) ocular disorders, such as age-related macular degeneration (AMD), diabetic retinopathy, glaucoma, and uveitis. The term “disorder” is used here interchangeably with “disease,” “condition,” and similar terms to refer to any health disorder or abnormal functional state of an organism, e.g., any condition for which medical and / or surgical intervention is indicated or for which the subject is appropriately required to undergo medical and / or surgical treatment. It should be understood that the listing of specific disorders within specific categories is for convenience and is not intended to limit the invention. It should be understood that a certain disorder may be appropriately illustrated in multiple categories.

[0045] Complement regulatory proteins (CRPs) are proteins involved in regulating complement activity. Complement regulatory proteins can downregulate complement activity, for example, by inhibiting complement activation or by inactivating or accelerating the breakdown of one or more activated complement proteins. Examples of complement regulatory proteins include C1 inhibitors, C4 binding proteins, clatherin, vitronectin, CFH, factor I, and cell-binding proteins CD46, CD55, CD59, CR1, CR2, and CR3.

[0046] As used herein, “isolation” means 1) separated from at least some of its components that normally coexist in nature; 2) manufactured or purified by human intervention; and / or 3) not naturally occurring, but present in an artificial environment, for example. In general, unless otherwise specified or clearly demonstrated, any substance, product, drug, composition, etc., may be considered “isolated” in some cases.

[0047] Here, the term “bond” used with two or more parts means that these parts are physically bonded or linked to each other to form a molecular structure that is sufficiently stable under the conditions under which the bond is formed, and preferably under the conditions under which the novel molecular structure is utilized, for example, under physiological conditions, so that these parts remain bonded. In one preferred embodiment of the present invention, the bond is a covalent bond. In other embodiments, the bond is a non-covalent bond. Parts can be bonded directly or indirectly. When two parts are directly bonded, they are covalently bonded to each other or are close enough that intermolecular forces between the two parts maintain the bond. When two parts are indirectly bonded, they are covalently or non-covalently bonded to a third part that maintains the bond between the two parts. Generally, when two parts are said to be bonded by a “bonding part” or “bonding part”, the bond between the two bonded parts is indirect, and typically each of the bonded parts is covalently bonded to the bonding part. Two parts may be bonded using a “linker”. A linker can be any suitable part that reacts with the substance to be bonded in a reasonable amount, for a reasonable amount of time, under conditions consistent with the stability of the substance (which may be partially protected as appropriate depending on the conditions). Typically, a linker contains at least two functional groups, one of which reacts with the first substance and the other with the second. It is recognized that after the linker has reacted with the substance to be bonded, the term “linker” may refer to the part of the resulting structure derived from the linker, or at least the part not included in the reacted functional groups. The bonding part may include parts that do not participate in bonding with the substance to be bonded, the main purpose of which may be to spatially separate these substances from each other. Such parts may be called “spacers.”

[0048] As used herein, “physiological conditions” refers to a set of conditions, such as temperature, salt concentration, and pH, that mimic at least some of those typically found in living subjects, e.g., mammalian subjects. In some embodiments, physiological conditions refer to an aqueous medium, e.g., a medium containing at least 90%, 95%, 96%, 97%, 97%, 99%, or approximately 100% water on a volume / volume basis. In some embodiments, other liquids, if present, do not substantially affect the protein secondary or tertiary structure. In some embodiments, physiological conditions mimic at least some of those found in body fluids, e.g., blood or extracellular fluid, e.g., interstitial fluid, of mammalian subjects. For example, a variety of physiological conditions useful in in vitro assays are known in this field. Generally, a medium under physiological conditions contains physiological concentrations of salt, e.g., sodium chloride. In one embodiment, a physiological concentration of salt refers to a concentration in the range of about 250 mOsm / L to about 350 mOsm / L, for example, in the range of about 275 mOsm / L to about 325 mOsm / L, for example, a concentration of about 300 mOsm / L. In one embodiment, physiological conditions are approximately isotonic with body fluids, e.g., blood or extracellular fluid, e.g., interstitial fluid. In one embodiment, physiological conditions include a pH in the range of about 6.5 to about 7.8, for example, in the range of about 7.0 to about 7.5. In one embodiment, the physiological medium includes a buffering agent that helps maintain the pH of the medium within the physiological range. In one embodiment, physiological conditions include conditions such that typical mammalian proteins, proteins typically found in body fluids such as blood or extracellular fluid, substantially maintain the secondary and, where applicable, tertiary structures they normally have in the body fluids in which they are found. In one embodiment, typically, the components of the physiological medium are substantially nontoxic to mammalian cells at the concentrations present in the physiological medium. A variety of physiological mediums (sometimes called “buffers”) are listed in various standard reference books, such as those cited above (e.g., Sambrook, et al., Protocols series). In one embodiment, the physiological temperature is in the range of approximately 25°C to approximately 38°C, for example, approximately 30°C to approximately 37°C, for example, 35°C to 37°C.

[0049] As used herein, “polypeptide” refers to a polymer of amino acids, which may sometimes contain one or more amino acid analogs. A protein is a molecule composed of one or more polypeptides. Peptides are relatively short polypeptides, typically about 2 to 60 amino acids long, for example, 8 to 40 amino acids long. The terms “protein,” “polypeptide,” and “peptide” can be used interchangeably. As used herein, polypeptides may include amino acids that are naturally found in proteins, amino acids that are not naturally found in proteins, and / or amino acid analogs that are not amino acids. As used herein, an amino acid “analog” may be another amino acid that is structurally similar to an amino acid, or a non-amino acid compound that is structurally similar to an amino acid. Numerous analogues of the 20 amino acids commonly found in proteins ("standard amino acids") are known in this field. One or more amino acids in a polypeptide can be modified, for example, by the addition of carbohydrate groups, phosphate groups, farnesyl groups, isofarnesyl groups, fatty acid groups, linkers, or other chemicals for bonding, functionalization, or other modifications. Some non-limiting suitable analogues and modification methods are described in WO2004026328 and are discussed later in this specification. Polypeptides may be acetylated at the N-terminus, for example, and C-terminus may be amidated.

[0050] As used herein, the term “purification” refers to a substance that has been separated from at least some or most of the components that were originally present or present before purification when it was synthesized. Generally, such purification involves human manual work. The purified product may be partially purified, substantially purified, or pure. Such a drug may be, for example, pure to at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or more than 99%. In some embodiments, nucleic acids, polypeptides, or small molecules are purified to constitute at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or more of the total nucleic acid, polypeptide, or small molecule substance present in the preparation, respectively. In some embodiments, organic substances, such as nucleic acids, polypeptides, or small molecules, are purified to constitute at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or more of the total organic substances present in the preparation. Purity can be determined, for example, based on dry weight, peak size of chromatographic output figures (GC, HPLC, etc.), molecular abundance, electrophoresis method, band intensity on gel, spectral data (e.g., NMR), elemental analysis, high-throughput sequencing, mass spectrometry, or any quantitative method accepted in the art. In some embodiments, water, buffers, ions, and / or small molecules (e.g., synthetic precursors such as nucleotides or amino acids) may be present in the purified preparation. The purified drug may be produced by separating it from other substances (e.g., other cellular substances) or by producing it in a manner that achieves the desired degree of purity. In one embodiment, “partial purification” of a molecule produced by a cell means that the molecule produced by the cell is no longer present in the cell, for example, the cell has been lysed and, optionally, at least some of the cellular material (e.g., cell wall, cell membrane, organelles) has been removed and / or the molecule has been separated or isolated from at least some of the same type of molecules (proteins, RNA, DNA, etc.) that were present in the lysate.

[0051] The terms “recombinant host cell,” “host cell,” and other such terms mean a prokaryotic or eukaryotic cell or prokaryotic cell line containing an exogenous nucleic acid (typically DNA), such as an expression vector containing the nucleic acid encoding the polypeptide of interest. Such terms are understood to include the offspring of the original cell into which the vector or other nucleic acid has been introduced. Suitable host cells include any cell routinely used in the art to express polynucleotides (for example, for the production of polypeptides encoded by such polynucleotides), including prokaryotes such as Escherichia coli or Escherichia; other bacteria such as Lactobacillus, Bacillus (e.g., Bacillus subtilis), Salmonella, Pseudomonas, Streptomyces, Staphylococcus, etc., and eukaryotes such as fungi such as yeast (e.g., Pichia (e.g., Pichia pastris), Clyberomyces (e.g., Clyberomyces lactis), Hansenula (e.g., Hansenula polymorpha), etc. Other examples of fungal cells include filamentous fungi, such as those of the genera Aspergillus, Neurospora, Fusarium, or Trichoderma, such as strains of Aspergillus oryzae, Aspergillus nidurans, or Aspergillus niger; insect cells (e.g., Sf9), plant cells, and animal cells, such as CHO, R1.1, BW, LM, African green monkey kidney cells (e.g., COS-1, COS-7, BSC-1, BSC-40, and BMT-10), and mammalian cells such as cultured human cells. Genetically modified (e.g., transgenic) cells in plants or animals are also included, in which recombinant polypeptides are produced in at least some of these cells. Polypeptides may be secreted into milk or obtained from plant matter. Exogenous nucleic acids may be stably maintained as episomes, such as plasmids, or integrated into the genome of at least some host cells, possibly after replication or reverse transcription. Terms such as "host cell" are also used to refer to cells or cell lines that can be used as recipients of foreign nucleic acids before nucleic acid production. "Recombinant polynucleotides" are polynucleotides that generally contain nucleic acid sequences that are not known to be directly bound to each other.For example, a nucleic acid sequence may exist in different genes or different species, or one or more of the sequence may be a variant of a naturally occurring sequence, or at least part of it may be an artificial sequence that is not homologous to a naturally occurring sequence. A “recombinant polypeptide” is generally a polypeptide that contains amino acid sequences that are produced and / or are not known to be directly bound to each other, at least partly by transcription and translation of an exogenous nucleic acid by a recombinant host cell or a cell-free in vitro expression system. In the latter case, the recombinant polypeptide may be called a “chimeric polypeptide.” The amino acid sequence of a chimeric polypeptide may, for example, have a substantial portion of its chain length existing in different genes or different species, or one or more of the sequence may be a variant of a naturally occurring sequence, or at least part of it may be an artificial sequence that is not identical to, or in some embodiments is not homologous to, a naturally occurring sequence. A chimeric polypeptide is understood to contain two or more polypeptides. For example, the first and second polypeptides A and B of a chimeric polypeptide may be directly bound (AB or BA) or separated by a third polypeptide portion C (ACB or BCA). In one embodiment, portion C represents a polypeptide linker, which may be, for example, multiple glycine and / or serine residues or a variety of other amino acids. In one embodiment, two or more polypeptides may be linked by a non-polypeptide linker. As used herein, “recombination” includes polypeptides produced by linking (e.g., chemical linkage, enzymatic linkage) of short recombinant polypeptides that may be produced in recombinant host cells, in one embodiment. In one embodiment, the recombinant polypeptide may include a signal sequence that directs the secretion of the polypeptide or a sequence that directs the polypeptide to a specific compartment or organelle. Suitable sequences are known in the art. A suitable sequence may be selected for the host cell type of interest (e.g., bacteria, fungi, mammals, plants, etc.). The signal sequence may, in one embodiment, be located at or near the N-terminus or C-terminus (e.g., up to 10-50 amino acids). In one embodiment, the polypeptide includes a tag. The tag may be useful for facilitating the detection and / or purification of the protein containing it.Examples of tags include polyhistidine tags (e.g., 6X-His tags), glutathione-S-transferase tags, maltose-binding proteins, NUS tags, SNUT tags, Strep tags, and epitope tags such as V5, HA, Myc, or FLAG. In some embodiments, the protease cleavage site is located in the region between the tag and the polypeptide, allowing the polypeptide to be separated from the tag upon exposure to the protease. In some embodiments, the polynucleotide encoding the recombinant polypeptide is at least partially codon-optimized for expression in the host cell of interest (e.g., bacteria, fungi, mammals, plants, etc.). In various correspondences, the tag may be located at or near the N-terminus or C-terminus of the polypeptide (e.g., within 10-50 amino acids). The recombinant polypeptide can be isolated, purified, etc., using any of a variety of methods. See, for example, Sambrook, Protocols series, or other standards. In one embodiment, the method of use may utilize a specific binding agent such as a tag or antibody, and may include, for example, dialysis (e.g., using a membrane with a specified pore size), chromatography, precipitation, gel purification, or affinity-based methods.

[0052] The "reactive functional groups" used here include olefins, acetylenes, alcohols, phenols, ethers, oxides, halides, aldehydes, ketones, carboxylic acids, esters, amides, cyanates, isocyanates, thiocyanates, isothiocyanates, amines, hydrazines, hydrazones, hydrazides, diazos, diazoniums, nitros, nitriles, mercaptans, sulfides, disulfides, sulfoxides, sulfones, sulfonic acids, sulfinic acids, acetals, ketals, anhydrides, and sulfur compounds. This includes, but is not limited to, phosphates, sulfenic acid, isonitrile, amidine, imide, imidate, nitrone, hydroxylamine, oxime, hydroxamic acid, thiohydroxamic acid, allene, orthoester, sulfite, enamine, inamine, urea, pseudourea, semicarbozide, carbodiimide, carbamate, imine, azide, azo compound, azoxy compound and nitroso compound, N-hydroxysuccinimide ester, maleimide, sulfhydryl, etc. Methods for producing each of these functional groups are well known in the art, and their application or modification for specific purposes is within the capabilities of those skilled in the art (e.g., Sandler and Karo, eds. ORGANIC FUNCTIONAL GROUP PREPARATIONS, Academic Press, San Diego, 1989 and Hermanson, G., Bioconjugate Techniques, 2 nd (See ed., Academic Press, San Diego, 2008).

[0053] "Specific binding" generally refers to the physical binding between a target polypeptide (or more specifically, a target molecule) and a binding molecule such as an antibody or a ligand. This binding is typically due to the presence of specific structural features of the target such as an antigenic determinant, epitope, binding pocket or cleft recognized by the binding molecule. For example, if an antibody is specific for epitope A, the presence of a polypeptide containing epitope A or the presence of free unlabeled A in a reaction containing both free labeled A and the binding molecule that binds to it will reduce the amount of labeled A that binds to the binding molecule. Specificity need not be absolute and it should be understood that it generally refers to the circumstances under which binding occurs. For example, it is well known in the art that many antibodies cross-react with other epitopes in addition to those present on the target molecule. Such cross-reactivity may be tolerated depending on the application in which the antibody is used. One of ordinary skill in the art can select an antibody or ligand having a sufficient degree of specificity to appropriately perform a particular application (e.g., for target molecule detection, for therapeutic purposes, etc.). It is also understood that specificity can be evaluated in terms of additional factors such as the affinity of the binding molecule for the target relative to its affinity for other targets, e.g., competitor substances. If the binding molecule exhibits high affinity for the target molecule that is desired to be detected and low affinity for non-target molecules, then the antibody is likely an acceptable reagent. Once the specificity of the binding molecule has been established in one or more circumstances, it can be used in other, preferably similar, circumstances without re-evaluating its specificity. In certain embodiments, the affinity (measured by the equilibrium dissociation constant, Kd) of the two molecules exhibiting specific binding is 10 -3 M or less, e.g., 10 -4 M or less, e.g., 10 -5 M or less, e.g., 10 -6 M or less, 10 -7 M or less, 10 -8 M or less or 10 -9 M or less under the conditions tested, e.g., under physiological conditions.

[0054] The “subjects” treated by the present invention are typically humans, non-human primates, or lower animals (e.g., mice or rats) that express or contain at least one primate (e.g., human) complement component C3, and optionally one or more further primate complement components. In some embodiments, the subject is male. In some embodiments, the subject is female. In some embodiments, the subject is an adult, e.g., a human at least 18 years old, e.g., 18–100 years old. In some embodiments, the human subject is at least 12 years old. In some embodiments, the subject is an adult, e.g., a human at least 18 years old, e.g., 18–100 years old. In some embodiments, the subject is at least 40, 45, 50, 55, 60, 65, 70, 75, or 80 years old. In some embodiments, the subject is a child, e.g., a human at 0–4 years old or 5–11 years old.

[0055] Here, “treatment” as used in relation to treatment of the subject means providing treatment, that is, providing some type of medical or surgical operation to the subject. Treatment may be provided for the resolution, reduction, halt, prevention, or reduction of the likelihood of a disease, or for the resolution, reduction, halt, prevention, or reduction of the likelihood of one or more symptoms or findings of a disease. “Prevention” means preventing the occurrence of a disease, symptoms, or disease findings in at least some individuals for at least a certain period of time. Treatment may involve administering a compound or composition to a subject after the onset of one or more symptoms or findings that are indicative of a disease, for example, for the resolution, reduction, reduction of severity, and / or halt or prevention of the disease, and / or for the resolution, reduction, reduction of severity, and / or prevention of one or more symptoms or findings of the disease. A compound or composition may be administered to a subject who has developed a disease or who has an increased risk of developing a disease compared to members of the general population. A compound or composition may be administered to a subject who has developed the disease and is at increased risk of developing one or more specific symptoms or findings of the disease or of disease exacerbation compared to other individuals diagnosed with the disease or compared to the subject's typical or average risk of such symptoms or findings or exacerbations. For example, the subject may be exposed to a “trigger” that increases the subject’s risk of experiencing an exacerbation (e.g., a temporarily increased risk). A compound or composition may be administered prophylactically, i.e., before the onset of any symptoms or findings of the disease. Typically, in this case, the subject is at increased risk of developing the disease compared to members of the general population, for example, age, sex, and / or other demographic variables which may be conformed.

[0056] A “vector” can be any of a variety of nucleic acid molecules, viruses, or parts thereof that can mediate the insertion, introduction, transport, etc., of a target nucleic acid between genetic environments or into cells. The target nucleic acid can be bound, for example, to a vector using restriction and ligation. A vector contains nucleic acids that can be encapsulated in, for example, DNA or RNA plasmids, cosmids, naturally occurring or modified viral genomes or parts thereof, viral capsids, minichromosomes, artificial chromosomes, etc. Plasmid vectors typically contain a replication origin (for example, for replication in prokaryotic cells). Plasmids contain part or all of a viral genome (for example, a viral promoter, enhancers, processing or packaging signals and / or a sequence sufficient to produce nucleic acids that can be integrated into the host cell genome and / or an infectious virus). A virus or part thereof that can be used to introduce nucleic acids into cells may be called a viral vector. Viral vectors include, for example, adenoviruses, adeno-associated viruses, retroviruses (e.g., lentiviruses, vaccinia viruses and other poxviruses), herpesviruses (e.g., herpes simplex virus), and others. Baculoviruses are, for example, for use in insect cells. A wide range of plant viral vectors are known, including, for example, those based on or containing cauliflower mosaic virus, tobacco mosaic virus, or one or more of their genetic elements (e.g., cauliflower mosaic virus 35S promoter). When introduced into a host cell, a viral vector may or may not contain sufficient viral genetic information for the production of an infectious virus; that is, a viral vector may be replicable or replication-deficient. In some embodiments, for example, when there is insufficient information for the production of an infectious virus, for example, if virus production is desired, it may be supplied by the host cell or another vector introduced into the cell. In some embodiments, for example, if virus production is not desired, such information is not supplied. The nucleic acid to be introduced may be incorporated into a naturally occurring or modified viral genome or a part thereof, or it may exist as another nucleic acid molecule within the viral capsid.A vector may contain one or more nucleic acids encoding markers suitable for identifying and / or selecting cells that have taken up the vector. Markers include, for example, various proteins that increase or decrease resistance or sensitivity to antibiotics or other drugs (e.g., proteins that contribute to antibiotic resistance, such as puromycin, hygromycin, or blastosidine), enzymes whose activity is detectable by assays known in the art (e.g., β-galactosidase or alkaline phosphatase), and proteins or RNAs that detectably affect the phenotype of the expressing cell (e.g., fluorescent proteins). Vectors often have one or more suitable sites for restriction enzymes, which can be used to facilitate the insertion of nucleic acids, e.g., nucleic acids to be expressed, into the vector. An expression vector is a vector into which the desired nucleic acid is inserted or can be inserted to operably bind to a regulatory element (also called a “regulatory sequence,” “expression regulatory element,” or “expression regulatory sequence”) and which can be expressed as an RNA transcript (e.g., mRNA that can be translated into a protein or non-coding RNA). An expression vector includes a control sequence, such as an expression regulatory sequence, sufficient to direct the transcription of the operablely bound nucleic acid under at least certain conditions, and other elements necessary for or assisting expression may be supplied, for example, by a host cell or an in vitro expression system. Such control sequences typically include a promoter and may include an enhancer sequence or an upstream activator sequence. In one embodiment, the vector may include a sequence encoding a 5' untranslated region and / or a 3' untranslated region, which may include a cleavage and / or polyadenylation signal. Generally, the control elements may be included in the vector before the insertion of the nucleic acid to be expressed, included in the insertion nucleic acid, or inserted into the vector after the insertion of the nucleic acid to be expressed. The nucleic acid and control elements used herein can be said to be “operably bound” if they are covalently bound such that the expression or transcription of the nucleic acid is under the influence or control of the control element. For example, a promoter region is operablely bound to the nucleic acid if the promoter region can influence the transcription of the nucleic acid.Those skilled in the art will understand that while the exact nature of regulatory sequences useful for gene expression varies by species or cell type, they may generally include sequences involved in transcription initiation, RNA processing, or translation initiation, as needed. The selection and design of appropriate vectors and regulatory elements are within the capabilities and discretion of those skilled in the art. For example, a person skilled in the art will select a promoter (or other regulatory sequence) suitable for expression in a desired species (e.g., prokaryotic (bacteria) or eukaryotic (e.g., fungi, plants, mammalian species) or cell type). The vector may include a suitable viral promoter from, for example, cytomegalovirus (CMV), retrovirus, simian virus (e.g., SV40), papillomavirus, herpesvirus or other viruses that infect mammalian cells, or a mammalian promoter derived from a gene such as EF1 alpha, ubiquitin (e.g., ubiquitin B or C), globin, actin, or phosphoglycerate kinase (PGK), or a composite promoter such as a CAG promoter (a combination of the CMV initial enhancer element and the chicken terta-actin promoter), which can direct expression in mammalian cells. In one embodiment, a human promoter may be used. In one embodiment, a promoter that normally directs transcription by eukaryotic RNA polymerase I ("pol A promoter such as “pol II promoter”, for example U6, H1, 7SK, or tRNA promoter or a functional variant thereof may be used. In one embodiment, a promoter that normally directs transcription by eukaryotic RNA polymerase II (“pol II promoter”) or a functional variant thereof may be used. In another embodiment, a promoter that normally directs transcription by eukaryotic RNA polymerase III (“pol III promoter”), for example a promoter for ribosomal RNA transcription (other than 5S rRNA) or a functional variant thereof may be used. Those skilled in the art will select a promoter suitable for directing the transcription of the desired sequence. Examples of expression vectors that can be used in mammalian cells include, for example, the pcDNA vector series, the pSV2 vector series, the pCMV vector series, the pRSV vector series, the pEF1 vector series, and Gateway® vectors.In one embodiment, a controllable (e.g., inducible or repressible) expression regulatory element, such as a controllable promoter, is used to regulate expression, for example, by activating or increasing or blocking or decreasing it. In one embodiment, the vector comprises a polynucleotide sequence encoding a polypeptide, where the polynucleotide sequence is positioned in a frame with a nucleic acid inserted into the vector so that an N-terminal or C-terminal fusion is formed. In one embodiment, the polypeptide encoded by the polynucleotide sequence may include a signal sequence (directing the secretion of a protein) or a sequence directing the expressed protein to a specific organelle or intracellular location such as the nucleus or mitochondria. In one embodiment, the polypeptide includes a tag. The tag may be useful for facilitating the detection and / or purification of the protein containing it. Examples of tags include polyhistidine tags (e.g., 6X-His tags), glutathione-S-transferase, maltose-binding protein, NUS tags, SNUT tags, Strep tags, epitope tags, such as V5, HA, Myc, or FLAG. In one embodiment, the protease cleavage site is located in the region between the protein encoded by the inserted nucleic acid and the polypeptide, allowing the polypeptide to be separated from the tag upon exposure to the protease. The vector can be introduced into host cells using methods known in the art. Those skilled in the art will select an appropriate method based, for example, the vector, cell type, etc. Suitable methods include, for example, calcium phosphate-mediated transfection; transfection using any of a variety of commercially available reagents, such as lipid-based or non-lipid-based transfections, e.g., FuGENE, lipofectamine, TurboFect; electroporation; and particulate gunshot. Such methods are described in detail in Sambrook, Protocols series, and other standards.

[0057] As used herein, the term “aliphatic” refers to a hydrocarbon moiety that may be linear (i.e., unbranched), branched, or cyclic (including condensed, cross-linked, and spiro-condensed polycyclic), and may be fully saturated or have one or more unsaturated units, but is not aromatic. Unless otherwise specified, an aliphatic group contains 1 to 30 carbon atoms. In one embodiment, an aliphatic group contains 1 to 10 carbon atoms. In another embodiment, an aliphatic group contains 1 to 8 carbon atoms. In yet another embodiment, an aliphatic group contains 1 to 6 carbon atoms, and in yet another embodiment, an aliphatic group contains 1 to 4 carbon atoms. Suitable aliphatic groups include, but are not limited to, linear or branched alkyl groups, alkenyl groups and alkynyl groups and hybrids thereof such as (cycloalkyl)alkyl, (cycloalkenyl)alkyl or (cycloalkyl)alkenyl.

[0058] As used herein, “alkyl” refers to saturated linear, branched, or cyclic hydroxyl molecules (and ranges and all combinations and subcombinations of specific numbers of carbon atoms within them) having about 1 to about 22 carbon atoms, with about 1 to about 12 or about 1 to about 7 carbon atoms being preferred in some embodiments of the present invention. Alkyl molecules include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, t-butyl, n-pentyl, cyclopentyl, isopentyl, neopentyl, n-hexyl, isohexyl, cyclohexyl, cyclooctyl, adamantyl, 3-methylpentyl, 2,2-dimethylbutyl, and 2,3-dimethylbutyl.

[0059] The term "halo" used here refers to F, Cl, Br, or I.

[0060] As used herein, “alkanoyl” refers to a linear or branched aliphatic acyclic residue having approximately 1 to 10 carbon atoms (and all combinations of range and specific carbon number, as well as lower combinations), for example, approximately 1 to 7 carbon atoms bonded by a single bond to a terminal C=O group (sometimes called an “acyl group”), and which may be substituted. Alkanoyl groups include, but are not limited to, formyl, acetyl, propionyl, butyryl, isobutyryl, pentanoyl, isopentanoyl, 2-methyl-butyryl, 2,2-dimethoxypropionyl, hexanoyl, heptanol, octanoyl, etc., and for the purposes of this invention, the formyl group is considered an alkanoyl group. “Lower alkanoyl” refers to a linear or branched aliphatic acyclic residue having approximately 1 to approximately 5 carbon atoms (and all combinations of range and specific carbon number, as well as lower combinations), which may be substituted. Such groups include, but are not limited to, formyl, acetyl, propionyl, butyryl, isobutyryl, pentanoyl, and isopentanoyl.

[0061] As used herein, “aryl” refers to monocyclic or bicyclic aromatic ring systems having approximately 5 to 14 carbon atoms (and all combinations of the range and specific numbers of carbon atoms within that range, as well as lower combinations), preferably approximately 6 to 10 carbon atoms, and optionally substituted. Non-limiting examples include, but are not limited to, phenyl and naphthyl.

[0062] As used herein, “aralkyl” refers to an alkyl radical having an aryl substituent and having about 6 to about 22 carbon atoms (and all combinations of the range and specific numbers of carbon atoms within that range, as well as lower combinations), where in some embodiments, the number of carbon atoms is preferably about 6 to about 12. The aralkyl group may optionally be substituted. Non-limiting examples include, for example, benzyl, naphthylmethyl, diphenylmethyl, triphenylmethyl, phenylethyl, and diphenylethyl.

[0063] The terms “alkoxy” and “alkoxyl” used herein refer to alkyl-O-groups that may be substituted, where alkyl is defined above. Examples of alkoxy and alkoxyl groups include methoxy, ethoxy, n-propoxy, i-propoxy, n-butoxy, and heptoxy.

[0064] The term "carboxyl" used here refers to the -C(=O)OH group. The term "alkoxycarbonyl" used here refers to a -C(=O)O-alkyl group, where alkyl is defined as described above.

[0065] The term "aloyl" used here refers to the -C(=O)-aryl group, where aryl is defined as above. Examples of alloyl groups include benzoyl and naphthoyl.

[0066] The term “cyclic ring system” refers to non-aromatic, partially unsaturated or fully saturated 3- to 10-membered ring systems, including bicyclic and tricyclic systems that may contain monocyclic rings of 3 to 8 atoms in size, as well as aromatic 5- or 6-membered aryl groups or aromatic heterocyclic groups fused with non-aromatic rings. Such heterocyclic rings include heterocyclic rings that independently have 1 to 3 heteroatoms selected from the group consisting of oxygen, sulfur, and nitrogen. In some embodiments, the term heterocyclic ring refers to a non-aromatic 5-, 6-, or 7-membered ring group or polycyclic group in which at least one ring atom is a heteroatom selected from the group consisting of O, S, and N, and includes, but is not limited to, bicyclic or tricyclic groups that contain a fused 6-membered ring having 1 to 3 heteroatoms independently selected from the group consisting of oxygen, sulfur, and nitrogen. In some embodiments, “ring system” refers to a cycloalkyl group, where cycloalkyl groups refer to groups having 3 to 10, for example, 4 to 7 carbon atoms. Cycloalkyl includes, but is not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl, which may be substituted as needed. In some embodiments, “ring system” refers to a cycloalkenyl or cycloalkynyl moiety, which may be substituted as needed.

[0067] Typically, a substituted chemical moiety contains one or more substituents that replace hydrogen. Examples of substituents include, for example, halo, alkyl, cycloalkyl, aralkyl, aryl, sulfidyl, hydroxyl (-OH), alkoxyl, cyano (-CN), carboxyl (-COOH), -C(=O)O-alkyl, aminocarbonyl (-C(=O)NH2), -N-substituted aminocarbonyl (-C(=O)NHR"), CF3, CF2CF3, etc. For the above substituents, the R'' can independently be, for example, H, alkyl, cycloalkyl, aryl, or aralkyl.

[0068] The term "L-amino acid" used herein refers to either naturally occurring levorotatory α-amino acids found in proteins or their alkyl esters. The term "D-amino acid" refers to dextrorotatory α-amino acids. Unless otherwise specified, all amino acids cited herein are L-amino acids.

[0069] The term "aromatic amino acid" used here refers to an amino acid that contains at least one aromatic ring; for example, aromatic amino acids contain an aryl group.

[0070] The “aromatic amino acid analog” used here is an amino acid analog containing at least one aromatic ring, for example, this one contains an aryl group.

[0071] II. Complement system To facilitate understanding of the present invention, and without any intention to limit it, this chapter outlines the complement and its activation pathway. For a more detailed explanation, see, for example, Kuby Immunology, 6. th ed., 2006; Paul, WE, Fundamental Immunology, Lippincott Williams & Wilkins; 6 th This is described in ed., 2008; and in Walport MJ., Complement. First of two parts. N Engl J Med., 344(14): 1058-66, 2001.

[0072] The complement system is part of the innate immune system that plays a vital role in defending the body from infectious pathogens. The complement system includes more than 30 serum and cellular proteins involved in three major pathways known as the classical pathway, the alternative pathway, and the lectin pathway. The classical pathway is typically initiated by the binding of an antigen-IgM or IgG antibody complex to C1 (although certain other activators can also initiate this pathway). Activated C1 cleaves C4 and C2 into C4a and C4b, and C2a and C2b. C4b and C2a combine to form C3 convertase, which cleaves C3 to form C3a and C3b. The binding of C3b to C3 convertase produces C5 convertase, which cleaves C5 into C5a and C5b. C3a, C4a, and C5a are anaphyllotoxins that mediate multiple responses in acute inflammatory responses. C3a and C5a are also chemotactic factors that attract immune system cells such as neutrophils.

[0073] Alternative pathways are initiated and amplified, for example, by microbial surfaces and various complex polysaccharides. In this pathway, hydrolysis of C3 to C3(H2O) occurs spontaneously at low levels, leading to binding with factor B. This is then cleaved by factor D to produce liquid-phase C3 convertase, which activates complement by cleaving C3 into C3a and C3b. C3b binds to targets such as the cell surface and forms a complex with factor B, which is later cleaved by factor D to produce C3 convertase. The surface-bound C3 convertase further cleaves and activates another C3 molecule, and C3b rapidly deposits near the activation site to form yet another C3 convertase, which in turn produces yet another C3b. This process creates a cycle of C3 cleavage and C3 convertase formation that greatly amplifies the reaction. C3 cleavage and binding of another C3b molecule to C3 convertase produce C5 convertase. The C3 and C5 convertases in this pathway are regulated by host cell molecules CR1, DAF, MCP, CD59, and fH. The mechanisms of action of these proteins include pro-activation disruption (i.e., the ability to dissociate convertases), their ability as cofactors in the degradation of C3b or C4b by factor I, or both. Normally, the presence of complement regulatory proteins on the host cell surface prevents significant complement activation at the cell surface.

[0074] The C5 convertases produced through both pathways degrade C5 to produce C5a and C5b. C5b then binds with C6, C7, and C8 to form C5b-8, which catalyzes the polymerization of C9 to form the C5b-9 membrane invasion complex (MAC). MACs invade the target cell membrane, causing cell lysis. Even small amounts of MAC on the cell membrane can lead to a variety of consequences other than cell death.

[0075] The lectin complement pathway is initiated by the binding of mannose-binding lectin (MBL) and MBL-associated serine protease (MASP) to carbohydrates. The MB1-1 gene (known as LMAN-1 in humans) encodes a type I intrinsic membrane protein located in the intermediate region between the endoplasmic reticulum and the Golgi apparatus. The MBL-2 gene encodes a soluble mannose-binding protein present in serum. In the human lectin pathway, MASP-1 and MASP-2 are involved in the degradation of C4 and C2 proteins, resulting in the C3 converter described above.

[0076] Complement activity is regulated by various mammalian proteins called complement regulatory proteins (CCPs) or complement activation regulator (RCA) proteins (U.S. Patent 6,897,290). These proteins differ in ligand specificity and complement inhibition mechanism. They accelerate the normal breakdown of convertases and / or function as cofactors of factor I, enzymatically cleaving C3b and / or C4b to further fragments. CCPs are characterized by the presence of multiple (usually 4 to 56) homologous motifs, approximately 50 to 70 amino acids in length, known as short consensus repeats (SCRs), complement regulatory protein (CCP) modules, or SUSHI domains, comprising a conserved motif with four disulfide-bonded cysteine ​​(two disulfide bonds), proline, tryptophan, and numerous hydrophobic residues. The CCP family includes complement receptor 1 (CR1; C3b:C4b receptor), complement receptor 2 (CR2), membrane cofactor protein (MCP; CD46), complement degradation factor (DAF), complement H factor (fH), and C4b-binding protein (C4bp). CD59 is a membrane-bound complement regulatory protein that is structurally unrelated to CCPs. Complement regulatory proteins typically restrict complement activation, which can otherwise occur in mammalian, e.g., human host cells and tissues. Thus, “self” cells are usually protected from the harmful effects that would follow if complement activation proceeded on these cells. Deficiencies or deficiencies of complement regulatory proteins are involved in the pathogenesis of various complement-mediated disorders, such as those described herein.

[0077] III. Compstatin Analogues Compstatin is a cyclic peptide that binds to C3 and inhibits complement activation. U.S. Patent 6,319,897 describes a peptide having the sequence Ile-[Cys-Val-Val-Gln-Asp-Trp-Gly-His-His-Arg-Cys]-Thr (SEQ ID NO: 1), where the disulfide bond between the two cysteine ​​molecules is enclosed in parentheses. The name "compstatin" is not used in U.S. Patent 6,319,897, but it is understood that it was subsequently adopted in scientific and patent literature (see, for example, Morikis, et al., Protein Sci., 7(3): 619-27, 1998) to refer to a peptide that has the same sequence as SEQ ID NO: 2 disclosed in U.S. Patent 6,319,897, but whose C-terminus is amidated as shown in Table 1 (SEQ ID NO: 8). Throughout this specification, the term “compstatin” is used consistently in this manner (i.e., when referring to Sequence ID No. 8). Compstatin analogs with higher complement inhibitory activity than compstatin have been developed. For example, WO2004 / 026328 (PCT / US2003 / 029653), Morikis, D., et al., Biochem Soc Trans. 32(Pt 1): 28-32, 2004, Mallik, B., et al., J. Med. Chem., 274-286, 2005; Katragadda, M., et al. J. Med. Chem., 49: 4616-4622, 2006; See WO2007062249(PCT / US2006 / 045539);WO2007044668(PCT / US2006 / 039397), WO / 2009 / 046198(PCT / US2008 / 078593);WO / 2010 / 127336(PCT / US2010 / 033345) and the discussion below.

[0078] Compstatin analogs may be acetylated or amidated at, for example, the N-terminus and / or C-terminus. For example, a compstatin analog may be acetylated at the N-terminus and amidated at the C-terminus. As used in the Art, “compstatin” as used herein and the activity of the compstatin analogs described herein compared to the activity of compstatin refer to compstatin amidated at the C-terminus (Mallik, 2005, supra).

[0079] Compstatin concatemers or multimers or their complement inhibitor analogs are also used in the present invention.

[0080] The term “compstatin analog” as used herein encompasses compstatin and all of its complement inhibitory analogs. The term “compstatin analog” also encompasses compstatin and other compounds designed or identified based on compstatin, whose complement inhibitory activity is at least 50% of the activity of compstatin as measured, for example, using any complement activation assay recognized in the art or an assay substantially identical or equivalent thereto. Specific suitable assays are described in U.S. Patent 6,319,897, WO2004 / 026328, Morikis, supra, Mallik, supra, Katragadda 2006, supra, WO2007062249 (PCT / US2006 / 045539); WO2007044668 (PCT / US2006 / 039397), WO / 2009 / 046198 (PCT / US2008 / 078593); and / or WO / 2010 / 127336 (PCT / US2010 / 033345). The assay may be an ELISA assay, for example, measuring erythrolysis mediated by an alternative or classical pathway. In one embodiment, the assay described in WO / 2010 / 135717 (PCT / US2010 / 035871) is used.

[0081] The activity of the compstatin analog is its IC 50IC can be expressed as (the concentration of a compound that inhibits complement activation by 50%), and as is recognized in the art, 50 A lower value indicates higher activity. The activity of a compstatin analog preferred for use in the present invention is at least comparable to that of compstatin. It should be noted that certain modifications known to reduce or counteract complement inhibitory activity can be explicitly excluded from any embodiment of the present invention. IC of compstatin 50 It has been measured as 12 μM using an alternative pathway-mediated erythropoiesis assay (WO2004 / 026328). The accurate IC25 is measured with a certain compstatin analog. 50 It should be understood that the values ​​will vary depending on the experimental conditions (e.g., serum concentration used in the assay). For example, IC of multiple different compounds under substantially identical conditions. 50 Comparative values ​​obtained from experiments determining the IC are useful. In one embodiment, the IC of a compstatin analog is useful. 50 Compstatin IC 50 The following applies: In one embodiment of the present invention, the activity of a compstatin analog is 2 to 99 times that of compstatin (i.e., the IC of the analog). 50 Compstatin IC 50 (2 to 99 times lower). For example, the activity may be 10 to 50 times greater than the activity of compstatin or 50 to 99 times greater than the activity of compstatin. In one embodiment of the present invention, the activity of the compstatin analog is 99 to 264 times greater than the activity of compstatin. For example, the activity may be 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, or 264 times greater than the activity of compstatin. In one embodiment, the activity is 250 to 300, 300 to 350, 350 to 400, or 400 to 500 times greater than the activity of compstatin. The present invention further aims to provide a compstatin analog having 500 to 1000 times or more activity than compstatin. In one embodiment, the IC of a compstatin analog 50The concentration is approximately 0.2 to 0.5 μM. In one embodiment, the IC of a compstatin analog is used. 50 The concentration is approximately 0.1 to 0.2 μM. In one embodiment, the IC of a compstatin analog is used. 50 The concentration is approximately 0.05 to 0.1 μM. In one embodiment, the IC of a compstatin analog is present. 50 The concentration is approximately 0.001 to 0.05 μM.

[0082] Compstatin K that binds to C3 d This can be measured using isothermal titration calorimetry (Katragadda, et al., J. Biol. Chem., 279(53), 54987-54995, 2004). As has been observed in the art, there is a correlation between the binding affinity of various compstatin analogs to C3 and their activity, and K d A lower K indicates higher binding affinity. A linear correlation between binding affinity and activity has been observed for certain analogues tested (Katragadda, 2004, supra; Katragadda 2006, supra). In one embodiment of the present invention, the compstatin analogue is 0.1-1.0 μM, 0.05-0.1 μM, 0.025-0.05 μM, 0.015-0.025 μM, 0.01-0.015 μM, or 0.001-0.01 μM of K. d Then it is joined with C3.

[0083] Compounds “designed or identified based on compstatin” include, but are not limited to, compounds containing amino acid chains whose sequences are obtained by (i) modifying the sequence of compstatin (e.g., replacing one or more amino acids in the sequence of compstatin with different amino acids or amino acid analogs, inserting one or more amino acids or amino acid analogs into the sequence of compstatin, or removing one or more amino acids from the sequence of compstatin); (ii) randomizing one or more amino acids of compstatin and, optionally, selecting from a library of further modified phage display peptides according to method (i); or (iii) identifying compounds whose sequences are obtained by screening for compounds that compete with compstatin or any analog obtained by method (i) or (ii) with respect to binding to C3 or its fragments. Many useful compstatin analogs contain hydrophobic clusters, β-turns, and disulfide bridges.

[0084] In one embodiment of the present invention, the sequence of a compstatin analog includes, or substantially consists of, a sequence obtained by making one, two, three, or four substitutions to the sequence of compstatin, that is, by replacing one, two, three, or four amino acids in the sequence of compstatin with different standard or non-standard amino acids. In one embodiment of the present invention, the amino acid at position 4 is altered. In one embodiment of the present invention, the amino acid at position 9 is altered. In one embodiment of the present invention, the amino acids at positions 4 and 9 are altered. In one embodiment of the present invention, only the amino acids at positions 4 and 9 are altered. In one embodiment of the present invention, the amino acid at position 4 or 9 is altered, or in one embodiment, both the amino acids at positions 4 and 9 are altered, and two or fewer amino acids at positions selected from positions 1, 7, 10, 11, and 13 are also altered. In one embodiment of the present invention, the amino acids at positions 4, 7, and 9 are altered. In one embodiment of the present invention, the amino acids at positions 2, 12, or both are altered, but the alteration preserves the compound's ability to cyclize. Such changes at positions 2 and / or 12 may be in addition to changes at positions 1, 4, 7, 9, 10, 11 and / or 13. In some cases, any compstatin analog sequence obtained by substituting one or more amino acids in the compstatin sequence may contain up to one, two, or three additional amino acids at the C-terminus. In one embodiment, the additional amino acid is glycan. In some cases, any compstatin analog sequence obtained by substituting one or more amino acids in the compstatin sequence may further contain up to five or up to ten additional amino acids at the C-terminus. Unless otherwise noted or evident from the context, it should be understood that a compstatin analog may have one or more features or properties of any of the various embodiments described herein, and that features or properties of any embodiment may further characterize any other embodiment described herein.In one embodiment of the present invention, the sequence of a compstatin analog comprises or substantially comprises the same sequence as compstatin, except with respect to positions corresponding to positions 4 and 9 of the compstatin sequence.

[0085] Compstatin and certain compstatin analogs with somewhat superior activity contain only standard amino acids ("standard amino acids" are glycine, leucine, isoleucine, valine, alanine, phenylalanine, tyrosine, tryptophan, aspartic acid, asparagine, glutamic acid, glutamine, cysteine, methionine, arginine, lysine, proline, serine, threonine, and histidine). Certain compstatin analogs with improved activity incorporate one or more non-standard amino acids. Useful non-standard amino acids include mono- and polyhalogenated (e.g., fluorinated) amino acids, D-amino acids, homoamino acids, N-alkyl amino acids, dehydroamino acids, aromatic amino acids (except phenylalanine, tyrosine, and tryptophan), ortho-, meta- or para-aminobenzoic acids, phosphoamino acids, methoxylated amino acids, and α,α-disubstituted amino acids. In one embodiment of the present invention, a compstatin analog is designed by replacing one or more L-amino acids of a compstatin analog described elsewhere in this specification with the corresponding D-amino acids. Such compounds and methods of use are aspects of the present invention.Examples of useful non-standard amino acids include 2-naphthylalanine (2-NaI), 1-naphthylalanine (1-NaI), 2-indanylglycinecarboxylic acid (2Igl), dihydrotryptophan (Dht), 4-benzoyl-L-phenylalanine (Bpa), 2-α-aminobutyric acid (2-Abu), 3-α-aminobutyric acid (3-Abu), 4-α-aminobutyric acid (4-Abu), cyclohexylalanine (Cha), homocyclohexylalanine (hCha), 4-fluoro-L-tryptophan (4fW), 5-fluoro-L-tryptophan (5fW), 6-fluoro-L-tryptophan (6fW), It contains 4-hydroxy-L-tryptophan (4OH-W), 5-hydroxy-L-tryptophan (5OH-W), 6-hydroxy-L-tryptophan (6OH-W), 1-methyl-L-tryptophan (1MeW), 4-methyl-L-tryptophan (4MeW), 5-methyl-L-tryptophan (5MeW), 7-aza-L-tryptophan (7aW), α-methyl-L-tryptophan (αMeW), β-methyl-L-tryptophan (βMeW), N-methyl-L-tryptophan (NMeW), ornithine (orn), citrulline, norleucine, γ-glutamic acid, etc.

[0086] In one embodiment of the present invention, a compstatin analog comprises one or more Trp analogs (e.g., at positions 4 and / or 7 in the compstatin sequence). Examples of Trp analogs are described above. See also Beene, et. al. Biochemistry 41: 10262-10269, 2002 (which describes monohalogenated Trp analogs and polyhalogenated Trp analogs in particular); Babitzke & Yanofsky, J. Biol. Chem. 270: 12452-12456, 1995 (which describes methylated Trp, halogenated Trp and other Trp analogs and indole analogs in particular); and U.S. Patents 6,214,790, 6,169,057, 5,776,970, 4,870,097, 4,576,750 and 4,299,838. Other Trp analogs include variants in which the α or β carbon of the indole ring and optionally one or more positions (e.g., by methyl groups) are substituted. Amino acids containing two or more aromatic rings are included in the Trp analogs, including their substituted variants, unsubstituted variants, or variants substituted in other embodiments. In one embodiment of the present invention, for example, the Trp analog at position 4 is 5-methoxy, 5-methyl-, 1-methyl-, or 1-formyl-tryptophan. In one embodiment of the present invention, Trp analogs (e.g., analogs at position 4) containing 1-alkyl substituents, such as lower alkyl (e.g., C1-C5) substituents are used. In a particular embodiment, N(α)methyltryptophan or 5-methyltryptophan is used. In one embodiment, analogs containing 1-alkanoyl substituents, such as lower alkanoyl (e.g., C1-C5) are used. Examples include 1-acetyl-L-tryptophan and L-β-tryptophan.

[0087] In one embodiment, the Trp analog exhibits increased hydrophobicity compared to Trp. For example, the indole ring may be substituted with one or more alkyl (e.g., methyl) groups. In one embodiment, the Trp analog is involved in hydrophobic interactions with C3. Such a Trp analog may be located at position 4 in the sequence of compstatin, for example. In one embodiment, the Trp analog comprises a substituted or unsubstituted bicyclic aromatic ring component or two or more substituted or unsubstituted monocyclic aromatic ring components.

[0088] In one embodiment, the Trp analog exhibits an increased tendency to form hydrogen bonds with C3 compared to Trp, but without increased hydrophobicity. The Trp analog may have increased polarity compared to Trp and / or an increased ability to engage in electrostatic interactions with hydrogen bond donors on C3. Specific exemplary Trp analogs with increased hydrogen bond-forming properties include an electronegative substituent on the indole ring. Such a Trp analog may be located, for example, at position 7 in the sequence of compstatin.

[0089] In one embodiment of the present invention, the compstatin analog comprises one or more Ala analogs (e.g., at position 9 relative to the compstatin sequence) and one or more Ala analogs that are identical to Ala except that they contain one or more CH2 groups in their side chains. In one embodiment, the Ala analog is an unbranched monomethylamino acid such as 2-Abu. In one embodiment of the present invention, the compstatin analog comprises one or more Trp analogs (e.g., at positions 4 and / or 7 in the compstatin sequence) and an Ala analog (e.g., at position 9 in the compstatin sequence).

[0090] In one embodiment of the present invention, the compstatin analog is (X'aa) n -Gln-Asp-Xaa-Gly-(X”aa) mThe compound contains a peptide having the sequence (SEQ ID NO: 2), where each X'aa and each X”aa are independently selected from amino acids or amino acid analogs, Xaa is Trp or an analog of Trp, and n>1, m>1, and n+m is 5-21. The peptide has the core sequence Gln-Asp-Xaa-Gly, where Xaa is Trp or an analog of Trp, for example, an analog of Trp that has an increased tendency to form hydrogen bonds with H bond donors compared to Trp, but in one embodiment is not hydrophobic compared to Trp. For example, the analog is, The indole ring of Trp may be substituted with an electronegative group, such as a halogen such as fluorine. In one embodiment, Xaa is 5-fluorotryptophan. Unless otherwise proven, those skilled in the art will recognize that any non-natural peptide whose sequence contains this core sequence and inhibits complement activation and / or binds to C3 is designed based on the sequence of compstatin. In another embodiment, Xaa is an amino acid or an amino acid analog other than a Trp analog on which the Gln-Asp-Xaa-Gly peptide can form a β-turn.

[0091] In one embodiment of the present invention, the peptide has the core sequence X'aa-Gln-Asp-Xaa-Gly (SEQ ID NO: 3), where X'aa and Xaa are selected from Trp and Trp analogs. In one embodiment of the present invention, the peptide has the core sequence X'aa-Gln-Asp-Xaa-Gly (SEQ ID NO: 3), where X'aa and Xaa are selected from Trp, Trp analogs, and other amino acids or amino acid analogs containing at least one aromatic ring. In one embodiment of the present invention, the core sequence forms a β-turn in relation to the peptide. The β-turn is flexible and may allow the peptide to have two or more conformations, which can be evaluated, for example, using nuclear magnetic resonance (NMR). In one embodiment, X'aa is a Trp analog containing a substituted or unsubstituted bicyclic aromatic ring component or two or more substituted or unsubstituted monocyclic aromatic ring components. In one embodiment of the present invention, X'aa is selected from the group consisting of 2-naphthylalanine, 1-naphthylalanine, 2-indanylglycinecarboxylic acid, dihydrotryptophan, and benzoylphenylalanine. In one embodiment of the present invention, X'aa is an analog of Trp in which hydrophobicity is increased compared to Trp. For example, X'aa may be 1-methyltryptophan. In one embodiment of the present invention, Xaa is an analog of Trp in which the tendency to form hydrogen bonds is increased compared to Trp, but in one embodiment the hydrophobicity is not increased compared to Trp. In one embodiment of the present invention, the analog of Trp in which the tendency to form hydrogen bonds is increased compared to Trp includes modifications to the indole ring of Trp, such as substitution of the H atom at position 5 with a halogen atom. For example, Xaa may be 5-fluorotryptophan.

[0092] In one embodiment of the present invention, the peptide has the core sequence X'aa-Gln-Asp-Xaa-Gly-X”aa (SEQ ID NO: 4), where X'aa and Xaa are independently selected from Trp and Trp analogs, and X”aa is selected from His, Ala, Ala analogs, Phe analogs, and Trp analogs. In one embodiment of the present invention, X'aa is another Trp analog having an alkyl substituent on a 1-methyltryptophan or indole ring (e.g., at position 1, 4, 5, or 6) and exhibiting increased hydrophobicity compared to Trp. In one embodiment, X'aa is a Trp analog comprising a substituted or unsubstituted bicyclic aromatic ring component or two or more substituted or unsubstituted monocyclic aromatic ring components. In one embodiment of the present invention, X'aa is selected from the group consisting of 2-naphthylalanine, 1-naphthylalanine, 2-indanylglycinecarboxylic acid, dihydrotryptophan, and benzoylphenylalanine. In one embodiment of the present invention, Xaa is an analog of Trp that has an increased tendency to form hydrogen bonds with C3 compared to Trp, but in another embodiment, its hydrophobicity is not increased compared to Trp. In one embodiment of the present invention, the Trp analog that has an increased tendency to form hydrogen bonds compared to Trp includes modifications to the indole ring of Trp, such as substitution of the H atom at position 5 with a halogen atom. For example, Xaa may be 5-fluorotryptophan. In one embodiment, X”aa is an Ala analog such as Ala or Abu, or another unbranched monomethylamino acid. In one embodiment of the present invention, the peptide has the core sequence X'aa-Gln-Asp-Xaa-Gly-X”aa (SEQ ID NO: 4), where X'aa and Xaa are each independently selected from Trp, a Trp analog, and an amino acid or amino acid analog containing at least one aromatic side chain, and X”aa is selected from His, Ala, an Ala analog, Phe, and Trp. In one embodiment, X”aa is selected from a Trp analog, an aromatic amino acid, and an aromatic amino acid analog.

[0093] In one preferred embodiment of the present invention, the peptide is cyclic. The peptide has one side (X'aa)n The other is (X"aa) m Cyclization can occur through a bond between any two amino acids located within the molecule. In one embodiment, the cyclic portion of the peptide is 9 to 15 amino acids long, for example, 10 to 12 amino acids long. In another embodiment, the cyclic portion of the peptide is 11 amino acids long and has a bond (e.g., a disulfide bond) between the amino acids at positions 2 and 12. For example, the peptide may be 13 amino acids long, with a cyclic portion of 11 amino acids formed by a bond between the amino acids at positions 2 and 12.

[0094] In one embodiment, the peptide contains or comprises the sequence X'aa1-X'aa2-X'aa3-X'aa4-Gln-Asp-Xaa-Gly-X”aa1-X”aa2-X”aa3-X”aa4-X”aa5 (SEQ ID NO: 5). In one embodiment, X'aa4 and Xaa are selected from Trp and Trp analogs, and X'aa1, X'aa2, X'aa3, X”aa1, X”aa2, X”aa3, X”aa4 and X”aa5 are independently selected from amino acids and amino acid analogs. In one embodiment, X'aa4 and Xaa are selected from aromatic amino acids and aromatic amino acid analogs. One or more of X'aa1, X'aa2, X'aa3, X”aa1, X”aa2, X”aa3, X”aa4, and X”aa5 may correspond to the amino acids at the corresponding positions in compstatin. In one embodiment, X”aa1 is Ala or a monomethyl unbranched amino acid. The peptide may be cyclized by covalent bonds between (i) X'aa1, X'aa2, or X'aa3 and (ii) X”aa2, X”aa3, X”aa4, or X”aa5. In one embodiment, the peptide is cyclized by a covalent bond between X'aa2 and X”aa4. In one embodiment, the covalently bonded amino acids are each Cys, and the covalent bond is a disulfide (SS) bond. In another embodiment, the covalent bond is a CC, CO, CS, or CN bond. In one embodiment, one covalently bonded residue is an amino acid or amino acid analog having a side chain containing a primary or secondary amine, and the other covalently bonded residue is an amino acid or amino acid analog having a side chain containing a carboxylic acid group, and the covalent bond is an amide bond. Examples of amino acids or amino acid analogs having a side chain containing a primary or secondary amine include lysine and the general structure NH2(CH2) nThis includes diaminocarboxylic acids of CH(NH2)COOH, such as 2,3-diaminopropionic acid (dapa), 2,4-diaminobutyric acid (daba), and ornithine (orn) (wherein n=1(dapa), 2(daba), and 3(orn), respectively). Examples of amino acids having a side chain containing a carboxylic acid group include dicarboxyl amino acids such as glutamic acid and aspartic acid. Analogues such as β-hydroxy-L-glutamic acid may also be used. In one embodiment, the peptide is cyclized by a thioether bond, as described, for example, in PCT / US2011 / 052442 (WO2012 / 040259). For example, in one embodiment, a disulfide bond of any peptide is replaced with a thioether bond. In one embodiment, a cystathionine is formed. In one embodiment, the cystathionine is δ-cystathionine or γ-cystathionine. In one embodiment, the modification includes replacing the Cys-Cys disulfide bond between the cysteines X'aa2 and X”aa4 (or the corresponding position in the sequence) of SEQ ID NO: 5 by adding a CH2 to form a homocysteine ​​at X'aa2 or X”aa4, and introducing a thioether bond to form a cystathionine. In one embodiment, the cystathionine is γ-cystathionine. In another embodiment, the cystathionine is δ-cystathionine. Another modification according to the present invention includes replacing the disulfide bond with a thioether bond without adding a CH2 to form a lantithionine. In one embodiment, a compstatin analog having a thioether instead of a disulfide bond is more stable than a compstatin analog having a disulfide bond, at least under certain conditions.

[0095] In one embodiment, the compstatin analog is sequence Xaa1-Cys-Val-Xaa2-Gln-Asp-Xaa2 * -Gly-Xaa3-His-Arg-Cys-Xaa4 (Sequence ID 6) [During the ceremony, Xaa1 is Ile, Val, Leu, B 1-Ile, B 1 -Val, B 1 -Leu or Gly-Ile or B 1 -Gly-Ile is a dipeptide containing B 1 This represents the first blocking section; Xaa2 and Xaa2 * These are independently selected from Trp and its analogues; Xaa3 is His, Ala or an analogue of Ala, Phe, Trp or an analogue of Trp; Xaa4 is a dipeptide selected from L-Thr, D-Thr, Ile, Val, Gly, Thr-Ala, and Thr-Asn, or a tripeptide containing Thr-Ala-Asn, wherein any of the carboxyl-terminal -OH groups of L-Thr, D-Thr, Ile, Val, Gly, Ala, or Asn may form a second blockage B 2 It has been replaced by; The two Cys residues are linked by a disulfide bond. The compound contains a peptide having the following: In one embodiment, Xaa4 is a dipeptide or tripeptide Xaa5-Ala-Asn selected from Leu, Nle, His or Phe or Xaa5-Ala and Xaa5-Asn, where Xaa5 is selected from Leu, Nle, His or Phe, and any carboxy-terminal -OH of L-Thr, D-Thr, Ile, Val, Gly, Leu, Nle, His, Phe, Ala or Asn optionally forms a second blockage B 2 It has been replaced by: the two Cys residues are linked by a disulfide bond.

[0096] In other embodiments, Xaa1 is absent or is any amino acid or amino acid analog, Xaa2, Xaa2 * Xaa3 and Xaa4 are as defined above. If Xaa1 is absent, the N-terminal Cys residue is the blockade portion B that is bound to it. 1 It has.

[0097] In other embodiments, Xaa4 is any amino acid or amino acid analog, Xaa1, Xaa2, Xaa2 * And Xaa3 is as defined above. In other embodiments, Xaa4 is a dipeptide selected from the group consisting of Thr-Ala and Thr-Asn, where the carboxyl-terminal -OH or Ala or Asn optionally forms a second blockage B 2 It has been replaced by...

[0098] In any embodiment of the compstatin analog of Sequence ID No. 6, Xaa2 may be Trp.

[0099] In any embodiment of the compstatin analog of SEQ ID NO: 6, Xaa2 may be an analog of Trp comprising a substituted or unsubstituted bicyclic aromatic ring component or two or more substituted or unsubstituted monocyclic aromatic ring components. For example, Trp analogs may be selected from 2-naphthylalanine (2-NaI), 1-naphthylalanine (1-NaI), 2-indanylglycinecarboxylic acid (Igl), dihydrotryptophan (Dht), and 4-benzoyl-L-phenylalanine.

[0100] In any embodiment of the compstatin analog of SEQ ID NO: 6, Xaa2 may be an analog of Trp in which hydrophobicity is increased compared to Trp. For example, the analog of Trp may be selected from 1-methyltryptophan, 4-methyltryptophan, 5-methyltryptophan, and 6-methyltryptophan. In one embodiment, the analog of Trp is 1-methyltryptophan. In one embodiment, Xaa2 is 1-methyltryptophan, and Xaa2 * is Trp, Xaa3 is Ala, and the other amino acids are the same as those of compstatin.

[0101] In any embodiment of the compstatin analog of Sequence ID No. 6, Xaa2 *This could be an analog of Trp, for example, one that has an increased tendency to form hydrogen bonds with C3 compared to Trp, but in one embodiment is not hydrophobic compared to Trp. In one embodiment, the Trp analog contains an electronegative substituent on the indole ring. For example, the Trp analog may be selected from 5-fluorotryptophan and 6-fluorotryptophan.

[0102] In one embodiment of the present invention, Xaa2 is Trp, and Xaa2 * Xaa2 is a Trp analog that has an increased tendency to form hydrogen bonds with C3 compared to Trp, but in some embodiments, its hydrophobicity is not increased compared to Trp. In some embodiments of the compstatin analog of SEQ ID NO: 6, Xaa2 is a Trp analog that has increased hydrophobicity compared to Trp, such as a Trp analog selected from 1-methyltryptophan, 4-methyltryptophan, 5-methyltryptophan and 6-methyltryptophan, and Xaa2 * Xaa2 is an analog of Trp in which the tendency to form hydrogen bonds with C3 is increased compared to Trp, but in one embodiment, the hydrophobicity is not increased compared to Trp. For example, in one embodiment, Xaa2 is methyltryptophan, and Xaa2 * It is 5-fluorotryptophan.

[0103] In any of the above embodiments, Xaa3 is Ala. In any of the above embodiments, Xaa3 is a monomethyl unbranched amino acid, for example, Abu.

[0104] The present invention further provides a compstatin analog of the above-mentioned Sequence ID No. 6, wherein Xaa2 and Xaa2 * The present invention provides a compstatin analog in which Xaa3 is independently selected from Trp, Trp analogs, and other amino acids or amino acid analogs containing at least one aromatic ring, and Xaa3 is His, Ala or an Ala analog, Phe, Trp, a Trp analog, or another aromatic amino acid or aromatic amino acid analog.

[0105] In one embodiment of the present invention, the N-terminus or C-terminus of any of the compstatin analogs described herein is any portion that stabilizes the peptide against degradation that may occur in the blood or interstitial fluid of mammals (e.g., humans or non-human primates). For example, blockade portion B 1 This could be any part that alters the N-terminal structure of the peptide so as to inhibit the cleavage of the peptide bond between the N-terminal amino acid of the peptide and the adjacent amino acid. Blocking part B 2 This can be any part that alters the C-terminal structure of the peptide so as to inhibit the cleavage of the peptide bond between the C-terminal amino acid of the peptide and the adjacent amino acid. Any suitable blocking part known in the art can be used. In one embodiment of the present invention, blocking part B 1 It contains an acyl group (i.e., the portion remaining after removing the -OH group from a carboxylic acid). An acyl group typically contains 1 to 12 carbon atoms, for example, 1 to 6 carbon atoms. For example, in one embodiment of the present invention, the blocked portion B 1 This is selected from the group consisting of formyl, acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, etc. In one embodiment, the blocking portion B 1 is an acetyl group, that is, Xaa1 is Ac-Ile, Ac-Val, Ac-Leu, or Ac-Gly-Ile.

[0106] In one embodiment of the present invention, the blocking portion B 2 These are primary or secondary amines (where R is an organic part such as an alkyl group, such as -NH2 or -NHR). 1 )

[0107] In one embodiment of the present invention, the blocking portion B 1 This is a portion that neutralizes or reduces the negative charge that may be present at the N-terminus in physiological pH. In one embodiment of the present invention, the blocking portion B 2 This is either a part that neutralizes or reduces the negative charge that may be present at the C-terminus in physiological pH.

[0108] In one embodiment of the present invention, the compstatin analog is acetylated or amidated at the N-terminus and / or C-terminus, respectively. The compstatin analog may be acetylated at the N-terminus, or amidated at the C-terminus, or acetylated at the N-terminus and amidated at the C-terminus. In one embodiment of the present invention, the compstatin analog contains an alkyl or aryl group at the N-terminus instead of an acetyl group.

[0109] In one embodiment, the compstatin analog is sequence Xaa1-Cys-Val-Xaa2-Gln-Asp-Xaa2 * -Gly-Xaa3-His-Arg-Cys-Xaa4 (Sequence ID 7) [During the ceremony, Xaa1 is a dipeptide containing Ile, Val, Leu, Ac-Ile, Ac-Val, Ac-Leu, or Gly-Ile or Ac-Gly-Ile; Xaa2 and Xaa2 * These are independently selected from Trp and its analogues; Xaa3 is His, Ala or an analogue of Ala, Phe, Trp or an analogue of Trp; Xaa4 is a dipeptide selected from L-Thr, D-Thr, Ile, Val, Gly, Thr-Ala, and Thr-Asn, or a tripeptide containing Thr-Ala-Asn, wherein one of the carboxyl-terminal -OH groups of L-Thr, D-Thr, Ile, Val, Gly, Ala, or Asn is optionally replaced by -NH2; The two Cys residues are linked by a disulfide bond. The compound contains a peptide having the following: In one embodiment, Xaa4 is a dipeptide or tripeptide Xaa5-Ala-Asn selected from Leu, Nle, His or Phe or Xaa5-Ala and Xaa5-Asn, where Xaa5 is selected from Leu, Nle, His or Phe, and any carboxy-terminal -OH of L-Thr, D-Thr, Ile, Val, Gly, Leu, Nle, His, Phe, Ala or Asn optionally forms a second blockage B 2 It has been replaced by; The two Cys residues are linked by a disulfide bond.

[0110] One proposal, Xaa1, Xaa2, Xaa2 * Xaa3 and Xaa4 are as described above for the various embodiments of Sequence ID No. 6. For example, in one embodiment, Xaa2 * is Trp. In one embodiment, Xaa2 is an analog of Trp in which hydrophobicity is increased compared to Trp, such as 1-methyltryptophan. In one embodiment, Xaa3 is Ala. In one embodiment, Xaa3 is a monomethyl unbranched amino acid.

[0111] In one embodiment of the present invention, Xaa1 is Ile and Xaa4 is L-Thr.

[0112] In one embodiment of the present invention, Xaa1 is Ile, and Xaa2 * is Trp, and Xaa4 is L-Thr.

[0113] The present invention further comprises compstatin analogs of the above-mentioned Sequence ID No. 7, namely Xaa2 and Xaa2 * The present invention provides compstatin analogs in which Xaa3 is independently selected from Trp, Trp analogs, other amino acids or aromatic amino acid analogs, and Xaa3 is His, Ala or Ala analog, Phe, Trp, Trp analog, or another aromatic amino acid or aromatic amino acid analog.

[0114] In one embodiment of any of the compstatin analogs described herein, an analog of Phe is used instead of Phe.

[0115] Table 1 provides a non-limiting list of compstatin analogs useful in the present invention. Each analog is abbreviated in the left column, indicating specific modifications at designated positions (positions 1-13) compared to the parent peptide, compstatin. As used in the art, “compstatin” as used herein and the activity of the compstatin analogs described herein compared to the activity of compstatin refer to the C-terminally amidated compstatin peptide. Unless otherwise specified, the peptides in Table 1 are C-terminally amidated. Specific modifications are indicated in bold. The activity compared to compstatin is based on publicly available data and the assays described therein (WO2004 / 026328, WO2007044668, Mallik, 2005; Katragadda, 2006). When multiple publications reporting a single activity are referenced, the value from the most recently published publication was used, but it is recognized that values ​​may be adjusted if there are differences between assays. Furthermore, in certain embodiments of the present invention, when the peptides listed in Table 1 are used in the therapeutic compositions and methods of the present invention, it is understood that they are cyclized via a disulfide bond between two Cys residues. Other means of cyclizing peptides are also within the scope of the present invention. As described above, in various embodiments of the present invention, one or more amino acids of a compstatin analog (e.g., any of the compstatin analogs disclosed herein) may be N-alkyl amino acids (e.g., N-methyl amino acids). For example, but not limited to, at least one amino acid inside the cyclic portion of the peptide, at least one amino acid at the N-terminus of the cyclic portion and / or at least one amino acid at the C-terminus of the cyclic portion may be N-alkyl amino acids, such as N-methyl amino acids. In certain embodiments of the present invention, for example, a compstatin analog contains N-methylglycine at a position corresponding to position 8 of compstatin and / or position corresponding to position 13 of compstatin. In one embodiment, one or more compstatin analogs in Table 1 contain, for example, one or more N-methylglycine molecules at the position corresponding to position 8 and / or position 13 of compstatin.In one embodiment, one or more compstatin analogs in Table 1 contain, for example, at least one N-methylisoleucine at the position corresponding to position 13 of compstatin. For example, the C-terminus or near-C-terminus of the peptide sequence listed in Table 1 or any other compstatin analog sequence may be replaced with N-methylIle. As recognized, in one embodiment, the N-methylated amino acid contains N-methylGly at position 8 and N-methylIle at position 13. In one embodiment, the N-methylated amino acid contains N-methylGly in a core sequence such as SEQ ID NO: 3 or SEQ ID NO: 4. In one embodiment, the N-methylated amino acid contains N-methylGly in a core sequence such as SEQ ID NO: 5, SEQ ID NO: 6 or SEQ ID NO: 7.

[0116] [Table 1] NA=Not available

[0117] In one embodiment of the composition and method of the present invention, the compstatin analog has a sequence selected from sequences 9 to 36. In one embodiment of the composition and method of the present invention, the compstatin analog has a sequence selected from sequence numbers 14, 21, 28, 29, 32, 33, 34 and 36. In one embodiment of the composition and / or method of the present invention, the compstatin analog has a sequence selected from sequence numbers 30 and 31. In one embodiment of the composition and method of the present invention, the compstatin analog has the sequence of sequence number 28. In one embodiment of the composition and method of the present invention, the compstatin analog has the sequence of sequence number 32. In one embodiment of the composition and method of the present invention, the compstatin analog has the sequence of sequence number 34. In one embodiment of the composition and method of the present invention, the compstatin analog has the sequence of sequence number 36.

[0118] In one embodiment, the blocking portion B 1 This includes an amino acid that can be represented as Xaa0. In one embodiment, the blocking portion B 2This includes an amino acid that can be represented as XaaN. In one embodiment, the blocking portion B 1 and / or B 2 This includes non-standard amino acids such as D-amino acids and N-alkyl amino acids (e.g., N-methyl amino acids). In one embodiment, the blocking portion B 1 and / or B 2 This includes non-standard amino acids that are analogs of standard amino acids. In one embodiment, the amino acid analog includes a lower alkyl, lower alkoxy, or halogen substituent relative to the standard amino acid analog. In one embodiment, the substituent is on the side chain. In one embodiment, the substituent is on the alpha carbon atom. In one embodiment, the barrier portion B includes an amino acid, for example, a non-standard amino acid. 1 This is further divided into parts B 1a This includes, for example, the blocking portion B. 1 is B 1a -Xaa0 can be expressed as B 1a This can neutralize or reduce the positive charge that might otherwise be present at the N-terminus at physiological pH. In one embodiment, B 1a This includes, for example, an acyl group containing 1 to 12 carbon atoms, for example, 1 to 6 carbon atoms. In one embodiment, the blocking portion B 1a The blockage portion B is selected from the group consisting of formyl, acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, and the like. In one embodiment, the blockage portion B includes an amino acid, for example, a non-standard amino acid. 2 This is further divided into parts B 2a It may include, for example, the blocking portion B. 2 XaaN-B 2a It can be expressed as, where N represents the appropriate number of the amino acid (this is due to the numbering used in the peptide backbone). In one embodiment, B 2a This can neutralize or reduce any negative charge that might otherwise be present at the C-terminus at physiological pH. In one embodiment, B 2a This includes or consists of primary or secondary amines (e.g., NH2). Part B 1a -Xaa0 and / or XaaN-B 2aThe blocking activity is understood to be provided by the components of either or both of these portions in various embodiments. In one embodiment, a blocking portion or a part thereof, e.g., 1 amino acid residue, contributes to the affinity of the compound for C3 or C3b and / or improves the activity of the compound. In one embodiment, the contribution to the affinity or activity of the amino acid residue can be at least as important as the contribution to the blocking activity. For example, in one embodiment, B 1a -Xaa0 and / or XaaN-B 2a In Xaa0 and / or XaaN in can mainly function to increase the affinity or activity of the compound, but B 1a and / or B 2a can prevent the digestion of the peptide and / or the neutralization of the charge. In one embodiment, the compstatin analog has an amino acid sequence of any one of SEQ ID NOs: 5 to 36, where SEQ ID NOs: 5 to 36 are further extended at the N-terminus and / or C-terminus. In one embodiment, the sequence can be represented as B 1a -Xaa0-sequence-XaaN-B 2a where the sequence represents any one of SEQ ID NOs: 5 to 36, and B 1a and B 2a may or may not be present independently. For example, in one embodiment, the compstatin analog is B 1a -Xaa0-X’aa1-X’aa2-X’aa3-X’aa4-Gln-Asp-Xaa-Gly-X”aa1-X”aa2-X”aa3-X”aa4-X”aa5-XaaN-B 2a (SEQ ID NO: 37A), where X’aa1-X’aa2-X’aa3-X’aa4, Xaa, X”aa1, X”aa2, X”aa3, X”aa4 and X”aa5 are as shown in SEQ ID NO: 5 above.

[0119] In one embodiment, the compstatin analog is B 1a -Xaa0-Xaa1-Cys-Val-Xaa2-Gln-Asp-Xaa2 * -Gly-Xaa3-His-Arg-Cys-Xaa4-XaaN-B 2a(Sequence ID 38A) includes, where Xaa1, Xaa2, Xaa2 * Xaa3 and Xaa4 are as shown in Sequence ID 6 above, or Xaa1, Xaa2, Xaa2 * Xaa3 and Xaa4 are as shown in Sequence ID No. 6 or Sequence ID No. 7.

[0120] In one embodiment, the compstatin analog is B 1a -Xaa0-Xaa1-Xaa2-Xaa3-Xaa4-Xaa5-Xaa6-Xaa7-Xaa8-Xaa9-Xaa10-Xaa11-Xaa12-Xaa13-XaaN-B 2a (Sequence ID 39A) is included, where Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, Xaa9, Xaa10, Xaa11, Xaa12 and Xaa13 are identical to the amino acids at positions 1-13 of any of Sequence IDs 9-36.

[0121] In some embodiments, Xaa0 and / or XaaN in any compstatin analog sequence comprises an amino acid having an aromatic ring with one or more alkyl substituents. In some embodiments, the alkyl substituent is a lower alkyl substituent. For example, in some embodiments, the alkyl substituent is a methyl group or an ethyl group. In some embodiments, the substituent is located at any position that does not disrupt the aromaticity of the compound. In some embodiments, the substituent is located at any position that does not disrupt the aromaticity of the ring to which the substituent is attached. In some embodiments, the substituent is located at position 1, 2, 3, 4, or 5. In some embodiments, Xaa0 comprises an O-methyl analog of tyrosine, 2-hydroxyphenylalanine, or 3-hydroxyphenylalanine. For the purposes of the present invention, a lowercase “m” following a three-letter amino acid abbreviation may be used to specifically indicate that the amino acid is an N-methylamino acid. For example, when the abbreviation “mGly” is used herein, it means N-methylglycine (sometimes called sarcosine or Sar). In one embodiment, Xaa0 is mgly, Tyr, Phe, Arg, Trp, Thr, Tyr(Me), Cha, mPhe, mVal, mle, mAla, DTyr, DPhe, DArg, DTrp, DThr, DTyr(Me), mPhe, mVal, mle, DAla, or DCha, or includes these. For example, in one embodiment, the compstatin analog is sequence B 1 -Ile-[Cys-Val-Trp(Me)-Gln-Asp-Trp-mGly-Ala-His-Arg-Cys]-mIle-B 2 (Sequence ID 40A) or B 1 -Ile-[Cys-Val-Trp(Me)-Gln-Asp-Trp-mGly-Ala-His-Arg-Cys]-mIle-B 2 The peptide contains (SEQ ID NO: 41A). Two Cys residues are linked by a disulfide bond in the active compound. In one embodiment, the peptide is acetylated at the N-terminus and / or amidated at the C-terminus. In one embodiment, as described above, B 1 is B 1a-Xaa0 includes and / or B 2 XaaN-B 2a This includes, for example, B 1 This includes or consists of Gly, mgly, Tyr, Phe, Arg, Trp, Thr, Tyr(Me), mPhe, mVal, mle, mAla, DTyr, DPhe, DTrp, DCha, DAla, B 2 It contains NH2, for example, the carboxyl terminal OH of the ml is substituted with NH2. In one embodiment, B 1 This includes or consists of mgly, Tyr, DTyr, or Tyr(Me), and B 2 It contains NH2, for example, the carboxyl-terminal OH of mIle is substituted with NH2. In one embodiment, Ile at position Xaa1 is replaced with Gly. The complement inhibitory activity and / or C3b binding parameters of selected compstatin analogs are described in WO / 2010 / 127336 (PCT / US2010 / 033345) and / or Qu, et al., Immunobiology (2012), doi: 10.1016 / j.imbio.2012.06.003.

[0122] In one embodiment, the blocking moiety or a portion thereof, for example, one amino acid residue, may contribute to an increased affinity of the compound for C3 or C3b and / or improve the compound's activity. In one embodiment, the contribution of an amino acid or amino acid analog to affinity or activity may be more significant than the blocking activity.

[0123] In one embodiment of the composition and method of the present invention, the compstatin analog has the sequence shown in Table 1, but as described above, the Ac group is replaced by a different blocking moiety B. 1 It is replaced by the -NH2 unit as described here in another blocking section B 2 It has been replaced by...

[0124] In one embodiment, the compstatin analog binds to substantially the same region of the human C3 β-chain as the region to which compstatin binds. In one embodiment, the compstatin analog is a compound that binds to the C-terminal fragment of the human C3 β-chain with a molecular weight of approximately 40 kDa to which compstatin binds (Soulika, AM, et al., Mol. Immunol., 35: 160, 1998; Soulika, AM, et al., Mol. Immunol. 43(12): 2023-9, 2006). In another embodiment, the compstatin analog is a compound that binds to the compstatin binding site determined by the compstatin-C3 structure, for example, by the crystal structure or 3D structure determined by NMR. In another embodiment, the compstatin analog is a compound that can replace compstatin in the compstatin-C3 structure and can form substantially the same intermolecular contact with C3 as compstatin. In one embodiment, the compstatin analog is a compound that binds to a binding site of a peptide having a sequence listed in Table 1 within the peptide-C3 structure, for example, within the crystal structure, such as SEQ ID NOs: 14, 21, 28, 29, 32, 33, 34 or 36, 37, 37A, 38A, 39A, 40A or 41A or other compstatin analog sequences listed herein. In one embodiment, the compstatin analog is a compound that binds to a binding site of a peptide having a sequence ID NOs: 30 or 31 within the peptide-C3 structure, for example, within the crystal structure. In one embodiment, the compstatin analog is a compound that can replace the peptides of SEQ ID NOs. 9-36, for example, peptides with SEQ ID NOs. 14, 21, 28, 29, 32, 33, 34 or 36, 37, 37A, 38A, 39A, 40A or 41A or other compstatin analog sequences described herein within the peptide-C3 structure, and is capable of forming an intermolecular contact with substantially the same C3 as that peptide. In one embodiment, the compstatin analog is a compound that can replace the peptide of SEQ ID NOs. 30 or 31 within the peptide-C3 structure, and is capable of forming an intermolecular contact with substantially the same C3 as that peptide.

[0125] Those skilled in the art can easily determine, using conventional experimental methods, whether a compstatin analog binds to the C-terminal fragment of the β-chain of C3. For example, those skilled in the art can synthesize photocrosslinkable compstatin analogs by incorporating a photocrosslinkable amino acid, such as p-benzoyl-L-phenylalanine (Bpa), at the C-terminus of the sequence in the compound (Soulika, AM, et al, supra). In some cases, additional amino acids, such as epitope tags like FLAG tags or HA tags, can be incorporated to facilitate detection of the compound, for example, by Western blotting. The compstatin analog is incubated with the fragment to initiate crosslinking. Colocalization of the compstatin analog and the C3 fragment indicates binding. Surface plasmon resonance may be used to determine whether the compstatin analog binds to the compstatin binding site or fragment of C3. Those skilled in the art can use molecular modeling software programs to predict whether a compound forms intermolecular contact with a C3 that is substantially the same as that of a peptide having the sequence of compstatin or any of the peptides listed in Table 1, for example, SEQ ID NOs: 14, 21, 28, 29, 32, 33, 34, or 36, or, in some embodiments, SEQ ID NOs: 30, 31, 37, 37A, 38A, 39A, 40A, or 41A, or any other compstatin analog sequence listed herein.

[0126] Compstatin analogs can be prepared by various known peptide synthesis methods in the art via amino acid residue condensation. For example, compstatin analogs can be prepared by expressing a suitable nucleic acid sequence encoding them in vitro or in living cells using methods known in the art, following conventional peptide synthesis methods. For example, peptides can be synthesized using standard solid-phase methods described in Malik, supra, Katragadda, supra, WO2004026328 and / or WO2007062249. Potentially reactive moieties, such as amino and carboxyl groups and reactive functional groups, can be protected and then deprotected using various protecting groups and methodologies known in the art. For example, “Protective Groups in Organic Synthesis”, 3 rd See (ed. Greene, TW and Wuts, PG, Eds., John Wiley & Sons, New York: 1999). The peptides can be purified using standard methods such as reversed-phase HPLC. If necessary, diastereomer peptides may be separated using known methods such as reversed-phase HPLC. If desired, the preparation may be lyophilized and then dissolved in a suitable solvent, such as water. The pH of the resulting solution can be adjusted to, for example, a physiological pH using a base such as NaOH. If necessary, the peptide preparation may be characterized by mass spectrometry to confirm the mass and / or disulfide bond formation. See, for example, Mallik, 2005, and Katragadda, 2006.

[0127] In one embodiment, a compstatin analog may be or may contain a cell-reactive compstatin analog. A cell-reactive compstatin analog is a compound comprising a compstatin analog moiety and a cell-reactive functional group that can react with a functional group exposed on the cell surface, for example, under physiological conditions, and form a covalent bond. Thus, the cell-reactive compstatin analog becomes covalently bonded to the cell. Without wishing to be bound to any particular theory, a cell-tethered compstatin analog protects the cell from complement-mediated damage, for example, by binding to C3 on the cell surface and / or around the cell (this may be in the form of C3(H2O)), inhibiting C3 cleavage and activation, and / or by binding to C3b, preventing its deposition in the cell or participation in the complement activation cascade. In one embodiment of the present invention, isolated cells are brought into ex vivo (out of body) contact with a cell-reactive compstatin analog. In one embodiment of the present invention, the cells are present in an isolated tissue or organ, for example, a tissue or organ to be transplanted into a subject. In one aspect of the present invention, cells are brought into contact with a cell-reactive compstatin analog by targeting the cell-reactive compstatin analog in vivo. The cell-reactive compstatin analog then covalently binds to the cells in vivo. In one aspect, the present invention protects cells, tissues, and / or organs from the adverse effects of complement activation for at least two weeks without requiring any treatment during that time.

[0128] In one embodiment, the present invention provides and / or utilizes a compstatin analog comprising a targeting moiety that non-covalently binds to a target molecule present on the surface of a cell or tissue, or to extracellular material not bound to a cell or tissue. Such a compstatin analog is hereafter referred to as a “targeted compstatin analog.” The target molecule is often a protein or carbohydrate bound to the cell membrane and exposed on the cell surface. The targeting moiety causes the compstatin analog to target cells, tissues, or locations sensitive to complement activation. In one embodiment of the present invention, isolated cells are brought into contact with the targeted compstatin analog ex vivo (out of the body). In one embodiment of the present invention, the cells are present in an isolated tissue or organ, e.g., a tissue or organ to be transplanted into a subject. In one embodiment of the present invention, the targeted compstatin analog is administered to the subject so that it non-covalently binds to cells, tissues, or extracellular material in vivo. In one embodiment, the present invention method protects cells, tissues, and / or organs from the harmful effects of complement activation for at least two weeks, without requiring any treatment during that time. In one embodiment, the targeted compstatin analog comprises both a targeting moiety and a cell-reactive moiety. The targeting moiety targets the compstatin analog, for example, by non-covalently binding to a specific cell type or to molecules on such cells. The cell-reactive moiety then covalently binds to the cell or extracellular material. In another embodiment, the targeted compstatin analog does not contain a cell-reactive moiety.

[0129] In one embodiment, the compstatin analog may be or may contain a long-acting compstatin analog, wherein the long-acting compstatin analog includes a portion such as polyethylene glycol (PEG) that extends the lifespan of the compound in the body (e.g., by reducing clearance from the blood). In one embodiment, the long-acting compstatin analog does not include a targeting moiety or a cell-reactive moiety. In one embodiment, the long-acting compstatin analog includes a targeting moiety and / or a cell-reactive moiety.

[0130] Depending on the circumstances, a compstatin analog bound to a cell-reactive or targeting moiety may be modified by adding polyethylene glycol (PEG) or similar molecules to stabilize the compound, reduce its immunogenicity, extend its lifespan in the body, increase or decrease its solubility, and / or increase its resistance to degradation. Methods for PEGylation are well known in this art (Veronese, FM & Harris, Adv. Drug Deliv. Rev. 54, 453-456, 2002; Davis, FF, Adv. Drug Deliv. Rev. 54, 457-458, 2002); Hinds, KD & Kim, SW Adv. Drug Deliv. Rev. 54, 505-530 (2002; Roberts, MJ, Bentley, MD & Harris, JM Adv. Drug Deliv. Rev. 54, 459-476; 2002); Wang, YS et al. Adv. Drug Deliv. Rev. 54, 547-570, 2002). A wide variety of polymers, including modified PEGs such as PEGs and derivatized PEGs, to which polypeptides can be appropriately conjugated, are described in the Nektar Advanced Pegylation 2005-2006 Product Catalog, Nektar Therapeutics, San Carlos, CA, which also provides details of appropriate conjugation methods. In another embodiment, a compstatin analog is fused with the Fc domain or a portion thereof of an immunoglobulin. In yet another embodiment, a compstatin analog is conjugated with an albumin moiety or an albumin-binding peptide. Thus, in one embodiment, a compstatin analog is modified with one or more polypeptide components or non-polypeptide components, for example, by pegyrating the compstatin analog or conjugating it with another moiety. In one embodiment, the component is neither the Fc domain nor a portion thereof of an immunoglobulin.Compstatin analogs can be obtained as a single molecular species, as a polymer potentially containing multiple different molecular species (e.g., multiple different analogs), or as part of a supramolecular complex.

[0131] In one embodiment, a compstatin analog is a polyvalent compound comprising multiple compstatin analog moieties covalently or noncovalently bonded to a polymer backbone or polymer scaffold. The compstatin analog moieties may be identical or different. In one embodiment of the present invention, the polyvalent compound comprises multiple examples or multiple copies of a single compstatin analog moiety. In another embodiment of the present invention, the polyvalent compound comprises one or more examples of two or more different compstatin analog moieties, e.g., three, four, five or more different compstatin analog moieties. In one embodiment of the present invention, the number of compstatin analog moieties ("n") is 2 to 6. In another embodiment of the present invention, n is 7 to 20. In another embodiment of the present invention, n is 20 to 100. In another embodiment, n is 100 and 1,000. In another embodiment of the present invention, n is 1,000 to 10,000. In another embodiment, n is 10,000 to 50,000. In another embodiment, n is between 50,000 and 100,000. In yet another embodiment, n is between 100,000 and 1,000,000.

[0132] The compstatin analog moiety may be directly bonded to the polymer scaffold, or it may be bonded via a bonding portion that connects the compstatin analog moiety and the polymer scaffold. The bonding portion can bond to a single compstatin analog moiety and a polymer scaffold. Alternatively, the bonding portion may have multiple compstatin analog moieties bonded to it, so that the bonding portion bonds multiple compstatin analog moieties to the polymer scaffold.

[0133] In one embodiment, the compstatin analog comprises an amino acid having a side chain containing a primary or secondary amine, such as a Lys residue. For example, a Lys residue or a sequence containing a Lys residue is added to the N-terminus and / or C-terminus of the compstatin analog. In one embodiment, the Lys residue is separated from the cyclic portion of the compstatin analog by a rigid or flexible spacer. The spacer may include, for example, a substituted or unsubstituted saturated or unsaturated alkyl chain, an oligo(ethylene glycol) chain, and / or other parts, such as those described in Section VI with respect to linkers. The chain length may be, for example, 2 to 20 carbon atoms. In another embodiment, the spacer is a peptide. The peptide spacer may be, for example, 1 to 20 amino acids in length, for example, 4 to 20 amino acids in length. A suitable spacer may include, for example, multiple Gly residues, Ser residues, or both, or consist of them. Optionally, the amino acid having a side chain containing a primary or secondary amine and / or at least one amino acid in the spacer is a D-amino acid. Any of the various polymer skeletons or polymer scaffolds can be used. For example, the polymer skeleton or polymer scaffold may be a polyamide, polysaccharide, polyacrylamide, polymethacrylic acid, polypeptide, polyethylene oxide, or dendrimer. Suitable methods and polymer skeletons are described, for example, in WO98 / 46270 (PCT / US98 / 07171) or WO98 / 47002 (PCT / US98 / 06963). In one embodiment, the polymer skeleton or polymer scaffold comprises several reactive functional groups such as carboxylic acid groups, anhydride groups, or succinimide groups. The polymer skeleton or polymer scaffold is reacted with a compstatin analog. In one embodiment, the compstatin analog comprises any of a number of different reactive functional groups such as carboxylic acid groups, anhydride groups, or succinimide groups, which are reacted with appropriate groups of the polymer skeleton. Alternatively, monomer units that can bond to each other to form a polymer skeleton or polymer scaffold are first reacted with a compstatin analog, and the resulting monomers are polymerized.In other embodiments, short chains are polymerized, functionalized, and then a mixture of short chains of different compositions is assembled into a long polymer.

[0134] IV. Compstatin mimetics The structure of Compstatin is known in the art, and the NMR structures of numerous Compstatin analogs with higher activity than Compstatin are also known (Malik, supra). Compstatin mimetics can be designed using the information on the structure.

[0135] In one embodiment, a compstatin mimetic is any compound that competes with compstatin or any compstatin analog (e.g., a compstatin analog of the sequences listed in Table 1) for binding to C3 or a fragment thereof (such as the 40 kD fragment of the β-chain to which compstatin binds). In certain embodiments, a compstatin mimetic has activity equal to or greater than that of compstatin. In certain embodiments, a compstatin mimetic is more stable than compstatin, orally available, or has high bioavailability. A compstatin mimetic can be a peptide, nucleic acid, or small molecule. In certain embodiments, a compstatin mimetic is a compound that binds to the binding site of compstatin determined in a compstatin-C3 structure, e.g., a crystal structure or a 3-D structure obtained from NMR experiments. In certain embodiments, a compstatin mimetic is a compound that can replace compstatin within a compstatin-C3 structure and form intermolecular contacts with substantially the same C3 as compstatin does. In certain embodiments, a compstatin mimetic is a compound that binds to the binding site of a peptide having a sequence listed in Table 1 within a peptide-C3 structure, e.g., SEQ ID NO: 14, 21, 28, 29, 32, 33, 34, or 36, or in certain embodiments, SEQ ID NO: 30 or 31 or other compstatin analog sequences. In certain embodiments, a compstatin mimetic is a compound that can replace a peptide having a sequence listed in Table 1 within a peptide-C3 structure, e.g., SEQ ID NO: 14, 21, 28, 29, 32, 33, 34, or 36, or in certain embodiments, SEQ ID NO: 30 or 31 or other compstatin analog sequences, and form intermolecular contacts with substantially the same C3 as that peptide does. In certain embodiments, a compstatin mimetic has a non-peptide backbone but has side chains arranged in a sequence designed based on the sequence of compstatin.

[0136] Those skilled in the art will understand that once a specific desired conformation of a short peptide is identified, methods for designing peptides or peptide mimes that conform to that conformation are well known. See, for example, GR Marshall (1993), Tetrahedron, 49: 3547-3558; Hruby and Nikiforovich (1991), in Molecular Conformation and Biological Interactions, P. Balaram & S. Ramasehan, eds., Indian Acad. of Sci., Bangalore, PP. 429-455), Eguchi M, Kahn M., Mini Rev Med Chem., 2(5): 447-62, 2002. Particularly in relation to the present invention, the design of peptide analogs can be further refined by taking into account the involvement of various side chains of amino acid residues in the action of functional groups or in conformational considerations, for example, as has been reported in the art with respect to compstatin and its analogues.

[0137] Those skilled in the art will understand that peptide mimes can serve just as well as peptides for the purpose of providing the specific skeletal conformation and side-chain functional groups necessary to bind to C3 and inhibit complement activation. Therefore, it is considered within the scope of the present invention to produce and utilize compounds that bind to C3 and inhibit complement by using any naturally occurring amino acid, amino acid derivative, amino acid analog, or non-amino acid molecule that can bind to form a suitable skeletal conformation. Here, non-peptide analogs or analogs containing both peptide and non-peptide components may be referred to as “peptide mimes” or “isoster mimes” to represent peptide substitutions or derivatives that have largely equivalent skeletal conformational properties and / or other functionalities to the exemplary peptides in terms of inhibiting complement activation. More generally, a compstatin mime is any compound in which the pharmacophore can be positioned similarly to its position in compstatin, even if it has a different skeleton.

[0138] The use of peptide mimes for developing high-affinity peptide analogs is well known in this art. By assuming rotational bindings nearly identical to those of amino acid residues in peptides, analogs containing non-amino acid portions can be analyzed using known techniques, particularly the Ramachandran plot, to confirm conformational motifs (Hruby & Nikiforovich 1991).

[0139] Those skilled in the art can easily establish appropriate screening assays to identify further compstatin mimes and select those with desired inhibitory activity. For example, compstatin or its analogues can be labeled (e.g., with radiolabeling or fluorescent labeling) and brought into contact with C3 in the presence of various concentrations of the test compound. The ability of the test compound to reduce the binding of the compstatin analog to C3 is evaluated. Test compounds that significantly reduce the binding of the compstatin analog to C3 are considered candidate compstatin mimes. For example, test compounds that reduce the steady-state concentration of the compstatin analog-C3 complex or reduce the formation rate of the compstatin analog-C3 complex by at least 25% or at least 50% are considered candidate compstatin mimes. Those skilled in the art will recognize that numerous variations of this screening assay can be used. Compounds to be screened include natural products, aptamer libraries, phage display libraries, and compound libraries synthesized using combinatorial chemistry. The present invention encompasses synthesizing a combinatorial library of compounds based on the above core sequence and screening the library to identify compstatin mimes. Using either of these methods, it is also possible to identify novel compstatin analogs with higher inhibitory activity than those previously tested. It is understood that compstatin mimes can be used in the cell-reactive compounds of the present invention and that the present invention provides such cell-reactive compstatin mimes.

[0140] V. Cell-reactive or long-acting compstatin analogs As described above, the present invention provides and / or utilizes various cell-reactive compstatin analogs. In some aspects, cell-reactive compstatin analogs include compounds of formula ALM, where A is a moiety containing a cell-reactive functional group J, L is an optionally present binding site, and M includes a compstatin analog moiety. The compstatin analog moiety may include any of the compstatin analogs described above in various embodiments. Formula ALM encompasses embodiments in which AL is located at the N-terminus of the compstatin analog moiety, embodiments in which AL is located at the C-terminus of the compstatin analog moiety, embodiments in which AL is attached to the amino acid side chains of the compstatin analog moiety, and embodiments in which the same or different ALs are located at both ends of M. When a particular compstatin analog exists as a compstatin analog moiety in a compound of formula ALM, it is understood that the functional group of the compstatin analog reacts with the functional group of L to form a covalent bond with A or L. For example, a compstatin analog containing an amino acid having a side chain containing a primary amine (NH2) group (the compstatin analog is given by formula R 1 Cell-reactive compstatin analogs containing (which can be represented as -(NH2)) have a new covalent bond with L (e.g., NC) and hydrogen is lost, resulting in formula R 1It may have -NH-LA. Therefore, the term “compstatin analog moiety” encompasses molecular structures in which at least one atom of the compstatin analog is involved in a covalent bond with the second moiety (e.g., a modification of the side chain). The same applies to compstatin analog moieties present in the polyvalent compounds described above. In one embodiment, the blocking moiety at the N-terminus or C-terminus of a compstatin analog, e.g., the compstatin analog described in Chapter IV above, is replaced with AL in the structure of a cell-reactive compstatin analog. In one embodiment, A or L contains a blocking moiety. In one embodiment, a cell-reactive compstatin analog has at least about 10%, 20%, or 30% of the activity of a corresponding compstatin analog having the same amino acid sequence (and one or more blocking moieties, if applicable) but without the cell-reactive moiety, e.g., 30-40%, 30-50%, 30-60%, 30-70%, 30-80%, 30-90%, or more molar activity. In one embodiment, the cell-reactive compstatin analog comprises multiple compstatin analog moieties, and the molar activity of the cell-reactive compstatin analog is at least about 10%, 20%, or 30% of the total activity of the compstatin analog moieties, for example, 30-40%, 30-50%, 30-60%, 30-70%, 30-80%, 30-90%, or more.

[0141] The cell-reactive portion A may comprise any of several different cell-reactive functional groups J in various embodiments. Generally, cell-reactive functional groups can be selected based at least partially on factors such as: (a) a specific functional group to target; (b) the ability of the reactive functional group to react with the target functional group under physiologically acceptable ex vivo conditions (e.g., physiologically acceptable pH and molar osmotic pressure) and / or in vivo conditions (e.g., in blood); (c) the specificity of the reaction between the reactive functional group and the target functional group under physiologically acceptable ex vivo conditions and / or in vivo conditions; (d) the stability (e.g., under in vivo conditions) of the covalent bond that may result from the reaction between the reactive functional group and its target functional group; and (e) the ease of synthesis of cell-reactive compstatin analogs containing the reactive functional group. In some embodiments, a reactive functional group is selected that reacts with the target chemical group without dissociating a leaving group. In some embodiments, a reactive functional group is selected that causes dissociation of a leaving group upon reaction with the target. Compounds containing such groups may be useful, for example, for monitoring the progress and / or extent of a reaction. In one embodiment, the leaving group is physiologically acceptable to cells, tissues, or organs in the amount produced (e.g., based on the concentration and / or absolute amount produced) and / or medically acceptable to the subject in the amount produced (e.g., based on the concentration in the relevant bodily fluids such as blood and / or based on the absolute amount produced). In one embodiment, at least a portion of the leaving group produced ex vivo is removed, for example, by washing cells or washing or perfusing tissues or organs with saline solution.

[0142] In many embodiments, the cell-reactive functional group used in the present invention reacts with the side chain of an amino acid residue and / or the N-terminal amino group or C-terminal carboxyl group of a protein. In one embodiment, the cell-reactive functional group reacts with a sulfidyl (-SH) group present in the side chain of a cysteine ​​residue. In one embodiment, a maleimide group is used. The maleimide group reacts with the sulfidyl group of a cysteine ​​residue of a protein at physiological pH to form a stable thioether bond. In one embodiment, a haloacetyl group such as an iodoacetyl group or a bromoacetyl group is used. The haloacetyl group reacts with a sulfidyl group at physiological pH. The reaction of the iodoacetyl group proceeds by nucleophilic substitution between iodine and the sulfur atom of the sulfidyl group, producing a stable thioether bond. In another embodiment, an iodoacetamide group is used. In one embodiment, the cell-reactive functional group reacts with an amino (-NH2) group present in the N-terminus of a protein and an amino group (ε-amino group) present in the side chain of a lysine residue. In one embodiment, an active ester, such as succinimidyl ester (i.e., NHS ester), is used. For example, N-hydroxysuccinimide (NHS) or its water-soluble analog (sulfo-NHS) is used in the synthesis, and the resulting cell-reactive compstatin analog contains the NHS ester. In one embodiment, the cell-reactive functional group reacts with carboxyl (-COOH) groups present in the C-terminus of proteins and in the side chains of various amino acid residues. In another embodiment, the cell-reactive compstatin analog reacts with hydroxyl (-OH) groups present in the side chains of various amino acids and in the carbohydrate portion of glycosylated proteins.

[0143] Generally, the bond L may include one or more aliphatic and / or aromatic moieties that match the formation of a stable compound that bonds the part to be bonded. The term “stable” as used herein preferably means a compound that is stable enough to be manufactured and maintains the integrity of the compound for, for example, one or more periods useful for the purposes described herein. In some embodiments, L includes a saturated or unsaturated, substituted or unsubstituted, branched or unbranched aliphatic chain having a length of 1 to 30, 1 to 20, 1 to 10, 1 to 6, or 5 carbon atoms, where length refers to the number of C atoms in the main chain (longest chain). In some embodiments, the aliphatic chain includes one or more heteroatoms (O, N, S), these heteroatoms may be independently selected. In some embodiments, at least 50% of the atoms in the main chain of L are carbon atoms. In some embodiments, L is a saturated alkyl moiety (CH2) n The formula includes n, where n is between 1 and 30.

[0144] In one embodiment, L contains one or more heteroatoms and has a length with a total number of carbon atoms in the chain of 1 to 1000, 1 to 800, 1 to 600, 1 to 400, 1 to 300, 1 to 200, 1 to 100, 1 to 50, 1 to 30, or 1 to 10. In one embodiment, L is an oligo(ethylene glycol) moiety (-(O-CH2-CH2-) n ) includes, where n is 1-500, 1-400, 1-300, 1-200, 1-100, 10-200, 200-300, 100-200, 40-500, 30-500, 20-500, 10-500, 1-40, 1-30, 1-20, or 1-10.

[0145] In one embodiment, L includes an unsaturated moiety such as -CH=CH- or -CH2-CH=CH-; a moiety containing a non-aromatic ring system (e.g., a cyclohexyl moiety); an aromatic moiety (e.g., an aromatic ring system such as a phenyl moiety); an ether moiety (-COC-); an amide moiety (-C(=O)-N-); an ester moiety (-CO-O-); a carbonyl moiety (-C(=O)-); an imine moiety (-C=N-); a thioether moiety (-CSC-); an amino acid residue; and / or any moiety that can be formed by the reaction of two compatible reactive functional groups. In one embodiment, one or more portions of a binding or cell-reactive moiety are substituted by one or more hydrogen (or other) atoms on that portion independently replacing one or more portions containing, but not limited to, fatty; aromatic, aryl; alkyl, aralkyl, alkanoyl, aroyl, alkoxy; thio; F; Cl; Br; I; -NO2; -CN; -CF3; -CH2CF3; -CHCl2; -CH2OH; -CH2CH2OH; -CH2NH2; -CH2SO2CH3; - or -GRG1, where G is -O-, -S-, -NRG2-, -C(=O)-, -S(=O)-, -SO2-, -C(=O)O-, -C(=O)NRG2- -OC(=O)-, -NRG2C(=O)-, -OC(=O)O-, -OC(=O)NRG2-, -NRG2C(=O)O-, -NRG2C(=O)NRG2-, -C(=S)-, -C(=S)S-, -SC(=S)-, -SC(=S)S-, -C(=NRG2)-, -C(=NRG2)O-, -C(=NRG2)NRG3-, -OC(=NRG2)-, -NRG2C(=NRG3)-, -NRG2SO2-, -NRG2SO2NRG3- or -SO2NRG2-, where RG1, RG2 and RG3 each independently contain, but are not limited to, hydrogen, halogens, or optionally substituted aliphatic, aromatic, or aryl moieties. When a ring system exists as a substituent, it is understood that it may be bonded via a linear portion in some cases.The combinations of substituents and variable compounds intended by the present invention preferably result in stable compounds that are useful in one or more of the methods described herein, for example, for treating one or more of the disorders described herein and / or for contact with cells, tissues or organs, and / or for use as intermediates in the production of one or more such compounds.

[0146] In various embodiments, L may comprise one or more of the parts described in the preceding paragraph. In some embodiments, L comprises two or more distinct parts bonded together to form a structure having a length of typically 1 to about 60 atoms, 1 to about 50 atoms, for example, 1 to 40, 1 to 30, 1 to 20, 1 to 10, or 1 to 6 atoms, where length refers to the number of atoms in the main chain (longest chain). In some embodiments, L comprises two or more distinct parts bonded together to form a structure having typically 1 to about 40 carbon atoms in the main chain (longest chain), for example, 1 to 30, for example, 1 to 20, 1 to 10, or 1 to 6 carbon atoms. The structure of such a cell-reactive compstatin analog is generally represented by formula A-(L Pj ) can be expressed by jM, where j is typically between 1 and 10, and L Pj Each of these is independently selected from the parts described in the previous paragraph. In many embodiments, L comprises one or more carbon-containing chains such as -(CH2)n- and / or -(O-CH2-CH2-)n, which are covalently bonded to each other and / or to cell-reactive functional groups or compstatin analogs by moieties resulting from the reaction of two compatible reactive functional groups (e.g., amide, ester, or ether moieties). In some embodiments, L comprises an oligo(ethylene glycol) moiety and / or a saturated alkyl chain. In some embodiments, L is -(CH2) m -C(=O)-NH-(CH2CH2O) n (CH2) p C(=O)- or -(CH2) m -C(=O)-NH-(CH2) p (OCH2CH2) nContains C(=O)-. In one embodiment, m, n, and p are selected such that the number of carbon atoms in the chain is 1 to 500, for example, 2 to 400, 2 to 300, 2 to 200, 2 to 100, 2 to 50, 4 to 40, 6 to 30, or 8 to 20. In one embodiment, m is 2 to 10, n is 1 to 500, and / or p is 2 to 10. In one embodiment, m is 2 to 10, n is 1 to 400, and / or p is 2 to 10. In one embodiment, m is 2 to 10, n is 1 to 300, and / or p is 2 to 10. In one embodiment, m is 2 to 10, n is 1 to 200, and / or p is 2 to 10. In one embodiment, m is 2 to 10, n is 1 to 100, and / or p is 2 to 10. In one embodiment, m is 2 to 10, n is 1 to 50, and / or p is 2 to 10. In one embodiment, m is 2 to 10, n is 1 to 25, and / or p is 2 to 10. In one embodiment, m is 2 to 10, n is 1 to 8, and / or p is 2 to 10. In some cases, at least one -CH2- is replaced by CH-R, where R can be any substituent. In some cases, at least one -CH2- is replaced by a heteroatom, a cyclic system, an amide moiety, an ester moiety, or an ether moiety. In one embodiment, L does not contain alkyl groups with more than three carbon atoms in the longest chain. In one embodiment, L does not contain alkyl groups with four, five, six, seven, eight, nine, ten, or eleven carbon atoms in the longest chain.

[0147] In one embodiment of the present invention, A is a cell-reactive functional group J and a binding portion L P1 Linker L 1 It includes a reactive functional group that reacts with a compstatin analog to produce AM. In one embodiment, it includes two reactive functional groups and a bonding site L P2 Includes a bifunctional linker L 2 The reactive functional group of L reacts with appropriate reactive functional groups of A and M to produce the cell-reactive compstatin analog ALM. In one embodiment, the compstatin analog is formed at the binding site LP3 Linker L 3 This includes, for example, even if a linker containing a reactive functional group is present at the N-terminus or C-terminus, the portion containing the reactive functional group may be bonded to the N-terminus or C-terminus via the linker. Therefore, L is a plurality of binding sites L provided by, for example, A, a linker used to bond A and M, and / or a compstatin analog. P It may contain. If present within the structure ALM, L before the reaction 1 , L 2 , L 3 It is understood that certain reactive functional groups present in such structures undergo a reaction, and only a portion of the reactive functional group is present in the final structure ALM, and the compound includes a portion formed by the reaction of the functional group. Generally, when a compound contains two or more bonding sites, these bonding sites may be the same or different, and can be independently selected in various embodiments. Multiple bonding sites L P These can be linked together to form a larger junction L, and at least a portion of such junctions may have one or more compstatin analogs and / or cell-reactive functional groups bound to them. In molecules containing multiple compstatin analogs, the compstatin analogs may be identical or different, and if different, they may be independently selected. The same applies to junctions and reactive functional groups. The present invention encompasses the use of polyvalent compstatin analogs containing one or more cell-reactive functional groups and the use of concatemers of compstatin analogs containing one or more cell-reactive functional groups. In some embodiments, at least one linkage is a stable non-covalent bond, such as a biotin / (strept)avidin linkage or another non-covalent bond of substantially the same strength.

[0148] In one embodiment, the cell-reactive compstatin analog comprises a compstatin analog in which one of SEQ ID NOs: 3-36, 37A, 38A, 39A, 40A, or 41A is extended by one or more amino acids at the N-terminus, C-terminus, or both ends, and at least one of the amino acids has a side chain containing a reactive functional group such as a primary or secondary amine, a sulfidyl group, a carboxyl group (which may exist as a carboxylic acid group), a guanidino group, a phenol group, an indole ring, a thioether, or an imidazole ring. In one embodiment, the amino acids are L-amino acids. In one embodiment, one or more amino acids are D-amino acids. If multiple amino acids are added, the amino acids can be selected independently. In one embodiment, a reactive functional group (e.g., a primary or secondary amine) is used as a target for adding the molar containing the cell-reactive functional group. Amino acids having a side chain containing a primary or secondary amine include lysine (Lys) and the general structure NH2(CH2). nThe compounds include diaminocarboxylic acids of CH(NH2)COOH, such as 2,3-diaminopropionic acid (dapa), 2,4-diaminobutyric acid (daba), and ornithine (orn) (when n=1(dapa), 2(daba), and 3(orn), respectively). In one embodiment, at least one amino acid is cysteine, aspartic acid, glutamic acid, arginine, tyrosine, tryptophan, methionine, or histidine. Cysteine ​​has a side chain containing a sulfidyl group. Aspartic acid and glutamic acid have a side chain containing a carboxyl group (which becomes a carboxylic acid group when ionized). Arginine has a side chain containing a guanidino group. Tyrosine has a side chain containing a phenol group (which becomes a phenolate group when ionized). Tryptophan has a side chain containing, for example, an indole ring containing tryptophan. Methionine has a side chain containing, for example, a thioether group containing methionine. Histidine has a side chain containing an imidazole ring. A wide variety of non-standard amino acids are available, including naturally occurring and non-naturally occurring amino acids, that have side chains containing one or more such reactive functional groups. See, for example, Hughes, B. (ed.), Amino Acids, Peptides and Proteins in Organic Chemistry, Volumes 1-4, Wiley-VCH (2009-2011); Blaskovich, M., Handbook on Syntheses of Amino Acids General Routes to Amino Acids, Oxford University Press, 2010. The present invention encompasses embodiments in which one or more non-standard amino acids are used as targets for adding a moiety containing a cell-reactive functional group. If necessary, one or more amino acids may be protected during the synthesis of the compound. For example, one or more amino acids may be protected during a reaction that includes the target amino acid side chain. In one embodiment, a sulfidyl-containing amino acid is used as a target for adding a moiety containing a cell-reactive functional group, and the sulfidyl is protected while the compound is cyclized by forming an intramolecular disulfide bond with another amino acid such as cysteine.

[0149] This paragraph describes the invention using amino acids having side chains containing amine groups as examples. The present invention encompasses similar embodiments using amino acids having side chains containing different reactive functional groups. In one embodiment, an amino acid having a side chain containing a primary or secondary amine is directly bound via peptide bonds to the N-terminus or C-terminus of any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A. In another embodiment, an amino acid having a side chain containing a primary or secondary amine is bound to the N-terminus or C-terminus of any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A via a binding site that may contain one or more of the above binding sites. In another embodiment, at least two amino acids are attached to one or both ends. These two or more attached amino acids may be linked to each other by peptide bonds, or at least some of the attached amino acids may be linked to each other via a binding site that may contain one or more of the binding sites described herein. Therefore, in one embodiment, the cell-reactive compstatin analog comprises a compstatin analog moiety M of formula B1-R1-M1-R2-B2, where M1 represents any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A, R1 or R2 may be absent, and at least one of R1 and R2 contains an amino acid having a side chain containing a primary or secondary amine, and B1 and B2 are optionally present blocking moieties. R1 and / or R2 may be linked to M1 by a peptide bond or a non-peptide bond. R1 and / or R2 are linked to moiety L P3 It may include. For example, if R1 is formula M2-L P3 It has and / or R2 is formula L P3 -M2 may be present, in the formula, L P3 The binding site is M2, and M2 contains at least one amino acid having a side chain containing a primary or secondary amine. For example, M2 may be Lys or an amino acid chain containing Lys. In one embodiment, L P3It contains or consists of one or more amino acids. For example, L P3 The length is 1 to about 20 amino acids, for example, the length is 4 to 20 amino acids. In one embodiment, L P3 It contains or consists of multiple Gly, Ser, and / or Ala residues. In one embodiment, L P3 It does not contain amino acids containing reactive SH groups such as Cys. In one embodiment, L P3 It comprises an oligo(ethylene glycol) moiety and / or a saturated alkyl chain. In one embodiment, L P3 It is bonded to the N-terminal amino acid of M1 via an amide bond. In one embodiment, L P3 It is bonded to the C-terminal amino acid of M1 via an amide bond. The compound may be further extended at one or both ends by the addition of further binding sites and / or amino acids. The amino acids may be the same or different, and if different, they may be selected independently. In one embodiment, two or more amino acids having side chains containing reactive functional groups are used, and the reactive functional groups may be the same or different. Two or more reactive functional groups can be used as targets for the addition of two or more moieties. In one embodiment, two or more cell-reactive moieties are added. In one embodiment, a cell-reactive moiety and a targeted moiety are added. In one embodiment, the linker and / or cell-reactive moiety are attached to the amino acid side chain after the amino acids have been incorporated into the peptide chain. In one embodiment, in the synthesis of a cell-reactive compstatin analog, the linker and / or cell-reactive moiety are already attached to the amino acid side chain before the amino acids are used. For example, a Lys derivative with a linker attached to the side chain may be used. The linker may contain a cell-reactive functional group, or may be later modified to contain a cell-reactive functional group.

[0150] The following provides further details on specific cell-reactive compstatin analogs. In the following discussion, the amino acid sequence Ile-Cys is used as an example of the compstatin analog moiety. *-Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * A peptide containing -Thr(SEQ ID NO: 37) (corresponding to the compstatin analog of SEQ ID NO: 28, where the asterisk in the sequence of SEQ ID NO: 37 represents a cysteine ​​bonded by a disulfide bond in the active compound, and (1Me)Trp represents 1-methyl-tryptophan) is used; maleimide (abbreviation Mal) is used as an example of a cell-reactive functional group; and (CH2) is used as an example of a binding site. n and (O-CH2-CH2) n The invention uses; lysine as an example of an amino acid containing a reactive functional group (in some compounds); and acetylation and amidation of the N-terminus and C-terminus (represented as Ac and NH2 in italics, respectively) as examples of blockage sites that may be present in some compounds. It is understood that the compounds can be prepared using various synthetic methods and various precursors. The following descriptions of various synthetic methods and precursors are not intended to limit the invention. In general, any feature of any of the compounds described below can be freely combined with the features of other compounds described below, here, or elsewhere in this specification, and the invention encompasses such embodiments.

[0151] In one embodiment, the cell-reactive moiety is provided by a cell-reactive compound containing a maleimide group (as a cell-reactive functional group) and an alkanoic acid (RCOOH) (where R is an alkyl group). For example, 6-malemadecaproic acid (Mal-(CH2)5-COOH) shown below. [ka] You can use it.

[0152] In one embodiment, the cell-reactive moiety is provided by a derivative of an alkanoic acid in which the carboxylic acid moiety is activated, for example, the OH moiety is converted to a more preferred leaving group. For example, after reacting the carboxyl group of Compound I with EDC and then reacting with NHS (which may optionally be provided as water-soluble sulfo-NHS), an N-hydroxysuccinimide ester derivative of 6-maleimidocaproic acid, i.e., 6-maleimidohexanoic acid N-hydroxysuccinimide (NHS) ester (below)

Chemical Structure

[0153] Cell-reactive compstatin analogs can be obtained by modifying the N-terminus and / or C-terminus of the compound of SEQ ID NO: 37. For example, using Compound II, the following cell-reactive compstatin analog can be obtained by reaction with the N-terminal amino group of Ile. Maleimido-(CH2)5-C(=O)-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2 (SEQ ID NO: 38) In SEQ ID NO: 38, it is understood that the -C(=O) moiety is attached to the adjacent C-terminal amino acid (Ile) via a C-N bond (where N is part of the amino acid and not shown).

[0154] In other embodiments, the maleimide group is attached to the C-terminal Thr to obtain the following cell-reactive compstatin analog. Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-(C=O)-(CH2)5-maleimide (SEQ ID NO: 39)

[0155] In one embodiment, a cell-reactive compstatin analog can be synthesized using a bifunctional linker (e.g., a heterobifunctional linker). (CH2-CH2-O) n Part and (CH2) m An example of a heterobifunctional linker including a sub-sub [ka]

[0156] Compound III comprises a maleimide group as a cell-reactive functional group and an NHS ester moiety that readily reacts with an amino group (e.g., an N-terminal amino group or an amino group in an amino acid side chain).

[0157] Using an embodiment of compound III where n=2, the following cell-reactive compstatin analog can be obtained using the compstatin analog of SEQ ID NO: 37. Maleimide-(CH2)2-C(=O)-NH-CH2CH2OCH2CH2OCH2CH2C(=O)-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2 (SEQ ID NO: 40)

[0158] In the compound of Sequence ID No. 40, it is understood that the -C (=O) moiety is linked to the N-terminal amino acid (Ile residue) via a CN bond (wherein N is a part of an amino acid and is not shown). In one embodiment, the linker has the formula of compound III where n is 1 or greater: (CH2-CH2-O) n Examples of the values ​​of n in the part are listed here.

[0159] In one embodiment, compared to the compound of SEQ ID NO: 39 or SEQ ID NO: 40, the alkyl chain linking the maleimide moiety to the rest of the molecule contains more or fewer methylene units, the oligo(ethylene glycol) moiety contains more or fewer ethylene glycol units, and / or more or fewer methylene units are present adjacent to one or both ends of the oligo(ethylene glycol) moiety. Examples of cell-reactive compstatin analogs to illustrate such variants are shown below (SEQ ID NOs: 41-46).

[0160] Maleimide-(CH2)2-C(=O)-NH-CH2CH2OCH2CH2C(=O)-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2 (SEQ ID NO: 41) Maleimide-(CH2)3-C(=O)-NH-CH2CH2OCH2CH2OCH2C(=O)-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2 (SEQ ID NO: 42) Maleimide-(CH2)5-C(=O)-NH-CH2CH2OCH2CH2OCH2C(=O)-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2 (SEQ ID NO: 43) Maleimide-(CH2)4-C(=O)-NH-CH2CH2OCH2CH2OCH2CH2C(=O)-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2 (SEQ ID NO: 44) Maleimide-(CH2)2-C(=O)-NH-CH2CH2OCH2CH2OCH2CH2C(=O)-Ile-Cys *-Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2 (SEQ ID NO: 45) Maleimide-(CH2)5-C(=O)-NH-CH2CH2OCH2CH2OCH2C(=O)-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2 (SEQ ID NO: 46)

[0161] In one embodiment, SEQ ID NO: 37 is extended, for example with respect to the C-terminal bond, as shown below, to include a Lys residue at the N-terminus or C-terminus of the peptide. Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-Lys-NH2 (SEQ ID NO: 47)

[0162] In one embodiment, the Lys residue is bound to the N-terminus or C-terminus of SEQ ID NO: 37 via a peptide linker, as shown below for example regarding the C-terminal bond. Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-(Gly)5-Lys-NH2 (SEQ ID NO: 48)

[0163] In one embodiment, a linker containing a primary or secondary amine is added to the N-terminus or C-terminus of the compstatin analog. In another embodiment, the linker contains an alkyl chain and / or an oligo(ethylene glycol) moiety. For example, NH2(CH2CH2O) nCH2C(=O)OH (e.g., 8-amino-3,6-dioxaoctanoic acid (AEEAc) or 11-amino-3,6,9-trioxaundecanoic acid) or its NHS ester (e.g., NHS ester of 8-amino-3,6-dioxaoctanoic acid or 11-amino-3,6,9-trioxaundecanoic acid) can be used. In one embodiment, the resulting compound is as follows (wherein the portion provided by the linker is shown in bold): [ka] [ka]

[0164] In one embodiment, a Lys residue is attached to the N-terminus or C-terminus of SEQ ID NO: 37 via a linker containing a non-peptide moiety. For example, the linker may include an alkyl chain, an oligo(ethylene glycol) chain, and / or a cyclic system. In one embodiment, 8-AEEAc or its NHS ester can be used to obtain the following compound (wherein the portion provided by 8-AEEAc is shown in bold) (when Lys is attached at the C-terminus): [ka]

[0165] In Sequence IDs 49 and 50, it is understood that the -C(=O) portion is bound to the adjacent Ile residue via a CN bond (wherein N is a part of an amino acid and is not shown). Similarly, in Sequence ID 51, the -C(=O) portion is bound to the adjacent Lys residue via a CN bond (wherein N is a part of an amino acid and is not shown). Furthermore, in Sequence ID 51, it is understood that the NH portion is bound to the adjacent N-terminal amino acid (Thr) via a CN bond (wherein C is the carbonyl carbon of an amino acid and is not shown).

[0166] It is easy to obtain cell-reactive compstatin analogs by modifying the primary amine group of the compounds of SEQ ID NOs. 47-51. For example, the following cell-reactive compstatin analogs can be obtained by reacting the compounds of SEQ ID NOs. 47-51 (or other compounds containing a primary or secondary amine and a compstatin analog moiety) with 6-maleimidocaproic acid N-succinimidyl ester. Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-Lys-(C(=O)-(CH2)5-Mal)-NH2(Sequence ID 52) Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-(Gly)5-Lys--(C(=O)-(CH2)5-Mal)-NH2(Sequence ID 53) [ka] [ka] [ka]

[0167] In another embodiment, a cell-reactive compstatin analog is used in Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * It is represented as -Thr-Lys-C(=O)-CH2(OCH2CH2)2NH(C(=O)-(CH2)5-Mal)-NH2 (sequence number 57).

[0168] The present invention provides variants of sequence numbers 38-57, which are -Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys *-Thr- is replaced by an amino acid sequence containing any other compstatin analog, such as any of the amino acid sequences of SEQ ID NOs: 3-27 or 29-36, 37, 37A, 38A, 39A, 40A, or 41A, provided that the blocking moiety present at the N-terminus and / or C-terminus of the compstatin analog is absent, replaced by a linker (which may contain the blocking moiety), or bound to a different N-terminal or C-terminal amino acid present within the corresponding variant.

[0169] Other bifunctional crosslinking agents useful in various embodiments of the present invention, comprising maleimide as a cell-reactive moiety and an NHS ester as an amine-reactive moiety, include, for example, succinimidyl 4-(p-maleimidophenyl)butyrate (SMPB); succinimidyl 4-(N-maleimidomethyl)-cyclohexane-1-carboxylate (SMCC); and N-γ-maleimidobutyryl-oxysuccinimidone (GMBS). Adding a sulfonic acid to the NHS ring yields water-soluble analogs such as sulfo-succinimidyl(4-iodoacetyl)-aminobenzoate (sulfo-SIAB), sulfo-succinimidyl 4-(N-maleimidomethyl)-cyclohexane-1-carboxylate (sulfo-SMCC), sulfo-succinimidyl 4-(p-maleimidophenyl)butyrate (sulfo-SMPB), and sulfo-N-γ-maleimidobutyryl-oxysuccinimidone (sulfo-GMBS), eliminating the need for organic solvents. In one embodiment of the present invention, any of the above long-chain forms is used, which includes a spacer arm between the NHS ester portion and the rest of the molecule. The spacer may include, for example, an alkyl chain. An example is succinimidyl-4-[N-maleimidomethyl]cyclohexane-1-carboxy-[6-amidecaproate].

[0170] In one embodiment, a bifunctional linker is used that comprises an NHS ester (as an amine-reactive site) and an iodoacetyl group (which reacts with a sulfhydryl group). Such linkers include, for example, N-succinimidyl(4-iodoacetyl)-aminobenzoate (SIAB); succinimidyl 6-[(iodoacetyl)-amino]hexanoate (SIAX); succinimidyl 6-[6-(((iodoacetyl)amino)-hexanoyl)amino]hexanoate (SIAXX); succinimidyl 4-((iodoacetyl)amino)methyl)-cyclohexane-1-carboxylate (SIAC); and succinimidyl 6-((((4-(iodoacetyl)amino)methyl-cyclohexane-1-carbonyl)amino)hexanoate (SIACX).

[0171] In one embodiment, a bifunctional linker is used that comprises an NHS ester (as an amine-reactive moiety) and a pyrididysulfide group (as a cell-reactive moiety that reacts with a sulfidyl group). Examples include N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP); succinimidyloxycarbonyl-α-methyl-α-(2-pyridyldithio)toluene (SMPT), and types that include a sulfonic acid on the NHS ring and / or a spacer containing an alkyl chain between the NHS ester moiety and the rest of the molecule (e.g., succinimidyl 6-(3-[2-pyridyldithio]-propionamide)hexanoate) (LC-SPDP). Modifications of such linkers containing additional or different parts can be used. For example, a relatively long or short alkyl chain may be used in the spacer, or an oligo(ethylene glycol) moiety may be used instead of an alkyl chain.

[0172] In general, cell-reactive compstatin analogs can be synthesized using various methods. Cell-reactive compounds containing cell-reactive functional groups and linkers can often be purchased as pre-formed basic components. For example, 6-malemadecaproic acid and 6-maleimidocaproic acid N-hydroxysuccinimide are available from various suppliers. Alternatively, such compounds can be synthesized using methods known in the art. See, for example, Keller O, Rudinger J. Helv Chim Acta. 58(2): 531-41, 1975 and Hashida S, et al., J Appl Biochem., 6(1-2): 56-63, 1984. For further consideration of methods and reagents useful for conjugate synthesis, see Hermanson, G. supra and the references therein. The present invention generally encompasses any method for producing compounds containing a compstatin analog moiety and a cell-reactive functional group, and the compounds obtained thereby.

[0173] In one embodiment, amino acids having linkers attached to their side chains are used in the synthesis of linear peptides. Linear peptides can be synthesized using standard peptide synthesis methods known in the art, such as standard solid-phase peptide synthesis. The linear peptide is then cyclized (for example, by oxidizing a Cys residue to form an intramolecular disulfide). This cyclic compound can then be reacted with a linker containing a cell-reactive functional group. In another embodiment, the portion containing the cell-reactive functional group is reacted with the linear compound before cyclization. Generally, during the synthesis of cell-reactive compstatin analogs, the reactive functional groups can be appropriately protected to prevent undesirable reactions between them. The cell-reactive functional group, any amino acid side chain, and / or one or both ends of the peptide can be protected during the reaction and then deprotected. For example, the SH group of a Cys residue and / or an SH-reactive moiety such as maleimide can be protected until after cyclization to prevent reactions between them. Reaction conditions are selected, at least in part, based on the need for specific reactive functional groups, to obtain a reasonable yield within a reasonable time. The temperature, pH, and reagent concentrations can be adjusted to obtain the desired degree or rate of the reaction. See, for example, Hermanson, supra. The desired product can be purified to remove, for example, unreacted compounds containing cell-reactive functional groups, unreacted compstatin analogs, linkers, products other than the desired cell-reactive compstatin analog produced in the reaction, and other substances present in the reaction mixture. Compositions and methods for producing cell-reactive compstatin analogs, as well as intermediates in their synthesis, are aspects of the present invention.

[0174] In one embodiment of the present invention, the above linker is used to produce a compstatin analog containing a portion such as a polyethylene glycol (PEG) chain or other polymer that stabilizes the compound, extends its lifespan in the body, increases its solubility, reduces its immunogenicity, and / or increases its resistance to degradation. Without limiting the present invention, such portions may be referred to herein as “clearance reduction portions” (CRMs), and compstatin analogs containing such portions may be referred to as “long-acting compstatin analogs” (LACAs). In one embodiment, when a long-acting compstatin analog is administered intravenously to humans or non-human primates at a dose of 10 mg / kg or at doses of approximately 1–3 mg / kg, 3–5 mg / kg, 5–10 mg / kg, for example, 7 mg / kg, it has an average plasma half-life of at least 1 day, for example, 1–3 days, 3–7 days, 7–14 days, or 14–28 days. In one embodiment, when a long-acting compstatin analog is administered subcutaneously to humans or non-human primates at doses such as approximately 1-3 mg / kg, 3-5 mg / kg, 5-10 mg / kg, or, for example, 7 mg / kg, it has an average plasma half-life of at least 1 day, for example, 1-3 days, 3-7 days, 7-14 days, or 14-28 days. In one embodiment, when a long-acting compstatin analog is administered intravenously to humans or non-human primates at doses of, for example, about 1-3 mg / kg, 3-5 mg / kg, or 5-10 mg / kg, for example, 7 mg / kg, it has a mean plasma half-life (e.g., terminal phase half-life) of about 4-10 days, 5-9 days, 5-8 days, 6-9 days, 7-9 days, or 8-9 days, for example, about 4 days, 4.5 days, 5 days, 5.5 days, 6 days, 6.5 days, 7 days, 7.5 days, 8 days, 8.5 days, 9 days, 9.5 days, or 10 days.In one embodiment, a long-acting compstatin analog has an average plasma half-life (e.g., terminal phase half-life) of approximately 4–10 days, 5–9 days, 5–8 days, 6–9 days, 7–9 days, or 8–9 days, for example, approximately 4 days, 4.5 days, 5 days, 5.5 days, 6 days, 6.5 days, 7 days, 7.5 days, 8 days, 8.5 days, 9 days, 9.5 days, or 10 days when administered subcutaneously to humans or non-human primates at doses such as approximately 1–3 mg / kg, 3–5 mg / kg, 5–10 mg / kg, or 7 mg / kg. In one embodiment, the long-acting compstatin analog is characterized by being widely absorbed from the injection site over a period of time after subcutaneous injection, achieving blood concentrations equivalent to those achieved with the same amount of compound administered intravenously, for example, approximately 1–2 days after administration or thereafter. In one embodiment, the blood concentrations at approximately 2, 3, 4, 5, 6, 7, 8 days or thereafter after subcutaneous administration are approximately 5%, 10%, 15%, 20%, or 25% of the blood concentrations achieved with the same amount of compound administered intravenously. See, for example, Figure 11, which shows the pharmacokinetics of the exemplary compounds described herein after intravenous and subcutaneous administration approximately 1–2 days after administration. In one embodiment, the mean plasma half-life of a long-acting compstatin analog after intravenous administration of 10 mg / kg to humans or non-human primates is extended by at least twofold, e.g., 2–5fold, 5–10fold, 10–50fold, or 50–100fold, or 100–150fold, or 150–200fold, compared to a corresponding compstatin analog having the same amino acid sequence (and, where applicable, one or more blocking moieties) but lacking a CRM. In various embodiments, it is understood that such an extension of half-life may also be observed after administration by other routes, such as subcutaneous administration, and / or after use of other doses, e.g., other doses described herein, e.g., 20 mg / kg.

[0175] As described above, in one embodiment, any of the compstatin analogs of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A is extended by one or more amino acids at the N-terminus, C-terminus, or both ends, and at least one of these amino acids has a side chain containing a reactive functional group such as a primary or secondary amine, a sulfidyl group, a carboxyl group (which may exist as a carboxylic acid group), a guanidino group, a phenol group, an indole ring, a thioether, or an imidazole ring, thereby promoting the binding of the CRM to the reactive functional group that links the compstatin analog. It is understood that the corresponding compstatin analog that does not contain CRM may lack one or more such amino acids that are present in the corresponding long-acting compstatin analog. Therefore, the corresponding compstatin analogs containing any of sequence numbers 3-36, 37, 37A, 38A, 39A, 40A, or 41A and lacking CRM are understood to have “the same amino acid sequence” as sequence numbers 3-36, 37, 37A, 38A, 39A, 40A, or 41A, respectively. For example, the corresponding compstatin analogs containing sequence numbers 14, 21, 28, 29, 32, 33, 34, or 3 and lacking CRM are understood to have “the same amino acid sequence” as sequence numbers 14, 21, 28, 29, 32, 33, 34, or 36, respectively.

[0176] In one embodiment, the plasma half-life refers to the terminal phase half-life after a single intravenous administration. In another embodiment, the plasma half-life refers to the terminal phase half-life after reaching a steady state following multiple intravenous administrations. In another embodiment, the long-acting compstatin analog is administered to primates as a single or multiple intravenous dose, and the plasma C max The plasma C of the corresponding compstatin analog that does not contain CRM max It reaches at least 5 times, for example, 5 to 50 times. In one embodiment, after a single intravenous administration of a long-acting compstatin analog to primates or after multiple intravenous administrations, the plasma C max The plasma C of the corresponding compstatin analog that does not contain CRM maxIt can reach 10 to 20 times that amount. In one embodiment, primates are humans.

[0177] In one embodiment, the primate is a non-human primate, such as a monkey like a crab-eating macaque or a rhesus macaque.

[0178] In one embodiment, a dose of 10 mg / kg or 20 mg / kg to humans or non-human primates results in at least a twofold reduction in renal clearance compared to the corresponding compstatin analog, e.g., 2–5, 5–10, 10–50, 50–100, 100–150, or 150–200 times. In various embodiments, it is understood that such a reduction in renal clearance may also be observed after administration via other routes, such as subcutaneous clearance, and / or after use of other doses, e.g., other doses described herein, e.g., 20 mg / kg.

[0179] The concentration of compstatin analogs can be measured in blood and / or urine samples using, for example, UV, HPLC, mass spectrometry (MS), antibodies against CRM, or a combination of such methods such as LC / MS or LC / MS / MS. Pharmacokinetic parameters such as half-life and clearance can be determined using methods known to those skilled in the art. For example, pharmacokinetic analysis can be performed using WinNonlin software v 5.2 (Pharsight Corporation, St. Louis, MO).

[0180] In one embodiment, CRM is stable for at least 24 hours under physiological conditions. In another embodiment, CRM is stable for at least 24 hours in mammalian blood, plasma, or serum, e.g., primates, e.g., humans or non-human primates (e.g., monkeys). In various embodiments, at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% of CRM molecules remain after incubation under physiological conditions for 24, 48, 72, 96, 120, 144, or 168 hours or more. In various embodiments, at least 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, and 99% or more of the CRM molecule remain intact after incubation in blood, plasma, or serum at 37°C for 48, 72, 96, 120, 144, and 168 hours or more. Incubation may be carried out using CRM at concentrations ranging from 1 μL / ml to approximately 100 mg / ml in various embodiments. Samples may be analyzed at various time points. Size or integrity may be assessed using, for example, chromatography (e.g., HPLC), mass spectrometry, Western blotting, or any other suitable method. Such stability properties may be conferred to the CRM-bound portion. In various embodiments, long-acting compstatin analogs containing CRM may possess any of the above stability properties. In one embodiment, completeness with respect to the long-acting compstatin analog means that the compstatin analog moiety remains bound to the CRM, and the CRM size remains approximately the same as at incubation or the start of administration.

[0181] In one embodiment, a long-acting compstatin analog has at least about 10%, 20%, or 30% of the molar activity of a corresponding compstatin analog having the same amino acid sequence (and, where applicable, one or more blocking moieties) but without CRM, e.g., 30-40%, 30-50%, 30-60%, 30-70%, 30-80%, 30-90%, or more. In one embodiment, the long-acting compstatin analog comprises multiple compstatin analog moieties, the molar activity of the long-acting compstatin analog is at least about 10%, 20%, or 30% of the total activity of the compstatin analog moieties, e.g., 30-40%, 30-50%, 30-60%, 30-70%, 30-80%, 30-90%, or more.

[0182] In one embodiment, polyethylene glycol (PEG) is (CH2CH2O) with a molecular weight of at least 500 daltons. n Includes a portion. In one embodiment, the linker is (CH2CH2O) with an average molecular weight of approximately 500;1,000;1,500;2,000;5,000;10,000;20,000;30,000;40,000;50,000;60,000;70,000;80,000;90,000;~100,000 Daltons. n Includes a portion.

[0183] In one embodiment, the average molecular weight of PEG is at least 20,000 daltons, and at most about 100,000; 120,000; 140,000; 160,000; 180,000; or 200,000 daltons. “Average molecular weight” refers to the number-average molecular weight. In one embodiment, (CH2CH2O) n The polyvariance D of the portion is between 1.0005 and 1.50, for example, 1.005 to 1.10, 1.15, 1.20, 1.25, 1.30, 1.40, or 1.50, or any value between 1.0005 and 1.50.

[0184] In one embodiment, (CH2CH2O) n The part is monodisperse, (CH2CH2O)n The polydispersity of the portion is 1.0. Such monodisperse (CH2CH2O) n The portion is publicly known in the art and is available for purchase from Quanta BioDesign (Powell, OH), and includes, in non-limiting examples, a monodisperse portion where n is 2, 4, 6, 8, 12, 16, 20, or 24.

[0185] In one embodiment, the chemical compound consists of multiple (CH2CH2O) n Including the portion, the (CH2CH2O) n The total molecular weight of the portion is approximately 1,000; 5,000; 10,000; 20,000; 30,000; 40,000; 50,000; 60,000; 70,000; 80,000; 90,000; ~100,000 Daltons. In one embodiment, the compound or (CH2CH2O) n The average total molecular weight of the portion is at least 20,000 Daltons, and at most about 100,000; 120,000; 140,000; 160,000; 180,000; or 200,000 Daltons. In one embodiment, the compound is (CH2CH2O) of a defined length, for example, n = 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28 or 30 or longer. n It contains multiple parts. In one embodiment, the compound is (CH2CH2O) of a defined length. n The part, the (CH2CH2O) n It contains a number of elements such that the total molecular weight of the elements is approximately 1,000; 5,000; 10,000; 20,000; 30,000; 40,000; 50,000; 60,000; 70,000; 80,000; 90,000; 100,000 Daltons. In one embodiment, the compound or (CH2CH2O) n The average total molecular weight of the portion is at least 20,000 daltons, and at most about 100,000; 120,000; 140,000; 160,000; 180,000; or 200,000 daltons. In one embodiment, n is about 30 to about 3000.

[0186] In one embodiment, the compstatin analog moiety is attached to each end of a linear PEG chain. For example, as described above, a bifunctional PEG having a reactive functional group at each end of the chain can be used. In one embodiment, the reactive functional groups are the same, but in another embodiment, different reactive functional groups are present at each end.

[0187] In one embodiment, multiple (CH2CH2O) n The portion is given in a branched structure. The branching may be attached to a linear polymer backbone (e.g., a comb-like structure) or may originate from one or more core groups (e.g., a star-like structure). In one embodiment, the branched molecule is (CH2CH2O) n It has 3 to 10 chains. In one embodiment, the branched molecule is (CH2CH2O) n It has 4 to 8 chains. In one embodiment, the branched molecule is (CH2CH2O) n It has 10, 9, 8, 7, 6, 5, 4, or 3 chains. In one embodiment, the star-shaped molecule is derived from a core group (CH2CH2O). n The chain has 10 to 100, 10 to 50, 10 to 30, or 10 to 20 strands. Therefore, in one embodiment, the long-acting compstatin analogs each have (CH2CH2O) via the functional group at the end of the chain. n The chain may contain, for example, 3 to 10, or for example, 4 to 8, compstatin analog moieties. In one embodiment, each long-acting compstatin analog is (CH2CH2O) via a functional group at the end of the chain. n The chain may contain 10 to 100 compstatin analog moieties. In one embodiment, the branched or star-shaped PEG branches (sometimes called “arms”) contain approximately equal numbers of (CH2CH2O) moieties. In one embodiment, at least some of the branches may be of different lengths. In one embodiment, one or more (CH2CH2O) nIt is understood that the chain does not have a compstatin analog moiety bound to it. In some embodiments, at least about 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 100% of the chain has a compstatin analog moiety bound to it.

[0188] In the group of compounds and compounds described herein, the polyethylene glycol moiety is depicted with the oxygen atom either to the right or to the left of the repeating unit. Even when only one orientation is depicted, the present invention also depicts the polyethylene glycol moiety of a compound or group of compounds in both orientations (i.e., (CH2CH2O) n and (OCH2CH2) n ) encompasses and, if a compound or group of compounds contains multiple polyethylene glycol moieties, any combination of orientations is included in this disclosure.

[0189] The following are some example formulas of monofunctional PEGs containing reactive functional groups. For illustrative purposes, the formulas shown include those containing NHS esters as reactive functional groups, but it is also possible to use other reactive functional groups, such as those shown above. In one embodiment, (CH2CH2O) n Although shown as ending with a methoxy group (OCH3) at the left end, it should be understood that chains shown below or elsewhere in this specification may end with different OR groups (e.g., an aliphatic group, an alkyl group, a lower alkyl group, or any other suitable PEG-terminated group) or OH groups. Also, in various embodiments, parts other than those shown may be (CH2CH2O) n It is also understood that the portion and the NHS group can be connected.

[0190] In one embodiment, monofunctional PEG is given by formula A [ka] It is a PEG, and in the formula, “Reactive functional group” and n are as defined above and as described in the Classes and subclasses herein; R 1is a hydrogen atom, an aliphatic atom, or any other suitable terminal group; T is covalent, or one or more carbon units of T may be independently -O-, -S-, -N(R) x )-, -C(O)-, -C(O)O-, -OC(O)-, -N(R x )C(O)-, -C(O)N(R x )-, -S(O)-, -S(O)2-, -N(R x )SO2- or SO2N(R x )- is replaced by C 1-12 It is a linear or branched hydrocarbon chain; R x Each of them independently represents hydrogen or C 1-6 It is an aliphatic.

[0191] An example of a monofunctional PEG of formula A is [ka] Includes.

[0192] In formula I, the part containing the reactive functional group is the general structure -CO-(CH2) where m is 2. m It has -COO-NHS. In one embodiment, the monofunctional PEG has the structure of formula I, where m is 1 to 10, for example 1 to 5. For example, in one embodiment, m is 3, as shown below. [ka]

[0193] [ka] In formula II, the portion containing the reactive functional group is the general structure -(CH2) where m is 1. m It has -COO-NHS. In one embodiment, the monofunctional PEG has the structure of formula II, where m is 1 to 10 (for example, m is 5 as shown in formula III below) or m is 0 (as shown in formula IIIa below). [ka] [ka]

[0194] In one embodiment, a difunctional linear PEG contains a moiety with a reactive functional group at both ends. The reactive functional groups may be the same (homodifunctional) or different (heterodifunctional). In one embodiment, the structure of the difunctional PEG may be symmetrical, in which the reactive functional group and -(CH2CH2O) n The oxygen atoms at each end of the chain are connected using the same region. In one embodiment, the two reactive functional groups and the PEG portion of the molecule are connected using different regions. An example structure of a bifunctional PEG is shown below. For illustrative purposes, the formula shown includes an NHS ester as the reactive functional group, but other reactive functional groups can also be used.

[0195] In one embodiment, a bifunctional linear PEG is of formula B [ka] The PEG is such that T and “reactive functional group” are each independently defined above and as described in the Classes and Subclasses herein, and n is defined above and as described in the Classes and Subclasses herein.

[0196] An example of a bifunctional PEG of formula B is [ka] Includes.

[0197] In formula IV, the part containing the reactive functional group is the general structure -(CH2) where m is 1. mIt has -COO-NHS. In one embodiment, the bifunctional PEG has the structure of formula IV, where m is 1 to 10, for example 1 to 5. In one embodiment, m is 0, for example, in one embodiment, the portion containing the reactive functional group has the general structure -COO-NHS. For example, in one embodiment, the bifunctional PEG has the structure of formula IVa shown below. [ka]

[0198] [ka]

[0199] In formula V, the part containing the reactive functional group is the general structure -CO-(CH2) where m is 2. m It has -COO-NHS. In one embodiment, the bifunctional PEG has the structure of formula V, where m is 1 to 10, for example 1 to 5. In one embodiment, for example, m is 2 as follows. [ka]

[0200] In one embodiment, the present invention provides a polymer-bonded compstatin analog. In one embodiment, the present invention provides PEG-containing compounds and compstatin analog conjugates of the group of compounds shown herein. In one embodiment, such conjugates are produced by reacting a functional group on a compstatin analog (e.g., an amine group, a hydroxyl group, or a thiol group) with a PEG-containing compound having the “reactive functional group” described herein. For example, formulas III and IV have the following structures, respectively [ka] or [ka] A compstatin analog conjugate having the following formula can be formed, in which, [ka] The symbol represents the binding site of the amine group on the compstatin analog. In one embodiment, the amine group is a lysine side chain group.

[0201] It is understood that any of the PEG-containing compounds and groups of compounds shown herein can be used to form a corresponding conjugate, depending on the selection of a reactive functional group and / or a compstatin functional group. For example, formulas IVa and Va can each form a compstatin analog conjugate having the following structure. [ka] [ka]

[0202] In one embodiment, the PEG elements of such a conjugate have an average molecular weight of about 20kD–100kD, about 20kD–90kD, about 20kD–80kD, about 20kD–70kD, about 20kD–60kD, about 20kD–50kD, about 30kD–80kD, about 30kD–70kD, about 30kD–60kD, about 30kD–50kD, about 30kD–45kD, about 35–50kD, about 35–45kD, about 36–44kD, about 37–43kD, about 38–42kD, or about 39–41kD. In one embodiment, the PEG elements of such a conjugate have an average molecular weight of about 40kD.

[0203] The term “difunctional” or “difunctionalization” is sometimes used to refer to a compound containing two compstatin analog moieties bonded to a CRM. Such a compound may be denoted by the letters “BF”. In some embodiments, the difunctional compound is symmetrical. In some embodiments, the bond between the CRM and each compstatin analog moiety of the difunctional compound is identical. In some embodiments, each bond between the CRM and each compstatin analog of the difunctional compound contains a carbamate. In some embodiments, each bond between the CRM and each compstatin analog of the difunctional compound contains a carbamate but does not contain an ester. In some embodiments, each compstatin analog of the difunctional compound is directly bonded to the CRM via a carbamate. In some embodiments, each compstatin analog of the difunctional compound is directly bonded to the CRM via a carbamate, and the difunctional compound has the following structure: [ka]

[0204] Embodiments and embodiments of the formulas described herein, [ka] is structure [ka] This represents the binding site of the lysine side chain group in a compstatin analog having the symbol "~", where the symbol "~" represents the binding site of the chemical moiety to the rest of the molecule or chemical formula.

[0205] In one embodiment, branched comb-shaped or star-shaped PEGs have multiple portions containing reactive functional groups, each containing -(CH2CH2O) n The reactive functional groups are included at the ends of the chain. The reactive functional groups may be the same, or there may be at least two different groups. In one embodiment, a branched comb-shaped or star-shaped PEG is given by the following formula [ka] [ka] [ka] [ka] [ka] [ka] It is a PEG, and in the formula, R 2 Each of these can be independently referred to as a “reactive functional group” or R 1 Thus, T, n, and “reactive functional group” are each independently defined above and as described in the Classes and Subclasses herein. An example structure of a branched PEG (having eight arms or branches) containing an NHS moiety as a reactive functional group is shown below.

[0206] [ka] [ka]

[0207] The structure of an example of a branched PEG (having four arms or branches) containing an NHS moiety as a reactive functional group is shown below. [ka] [ka]

[0208] The number of branches arising from the skeleton can vary. For example, the number of branches, 4, in formulas VI and VII above may be changed to any other integer from 0 to 10 in various embodiments. In one embodiment, one or more branches do not contain a reactive functional group, and these branches end with a -CH2CH2OH group or a -CH2CH2OR group as described above.

[0209] In one embodiment, the branched PEG has the structure of formula VII, VIII, or IX (or its variants with a different number of branches), where x is [ka] That is the case.

[0210] In one embodiment, the branched PEG has the structure of formula VII, VIII, or IX (or its variants with a different number of branches), where x is [ka] That is the case.

[0211] Naturally, the methylene (CH2) group in the above x portion can be replaced by a longer alkyl chain (CH2). m It may include (where m is 2, 3, 4, 5, 6, 8, 10, 20, or 30 or less), or it may include one or more of the other parts listed herein.

[0212] In one embodiment, an example of a branched PEG having an NHS reactive group or a maleimide reactive group is illustrated as follows. [ka] [ka] In one embodiment, a variant of formula X or XI is used in which three of the four branches, or each branch, contains a reactive functional group.

[0213] Another example of PEG is shown below. [ka] [ka]

[0214] As described above and as stated herein, in various embodiments, the peptide component of the long-acting compstatin analog and (CH2CH2O) n It is understood that any of the following moieties may be incorporated between the -R moiety: linear alkyl, ester, amide, aromatic ring (e.g., substituted or unsubstituted phenyl), substituted or unsubstituted cycloalkyl structure, or a combination thereof. In some embodiments, such a moiety increases the sensitivity of the compound to hydrolysis, thereby allowing the peptide portion of the compound to dissociate from the CRM. In some embodiments, such dissociation may increase the in vivo tissue penetration and / or activity of the compound. In some embodiments, the hydrolysis is a general (e.g., acid-base) hydrolysis. In some embodiments, the hydrolysis is enzyme-catalyzed, e.g., esterase-catalyzed. Naturally, both types of hydrolysis may occur. Examples of PEGs containing one or more such moieties and an NHS ester as a reactive functional group include: [ka] [ka] [ka]

[0215] In one embodiment, branched (multi-armed) PEG or star-shaped PEG comprises a pentaerythritol nucleus, a hexaglycerin nucleus, or a tripentaerythritol nucleus. In one embodiment, it is understood that not all branches originate from a single point.

[0216] Monofunctional, bifunctional, branched, and other PEGs containing one or more reactive functional groups can, in some embodiments, be obtained, for example, from NOF America Corp. White Plains, NY or BOC Sciences 45-16 Ramsey Road Shirley, NY 11967, USA, or prepared using methods known in the art.

[0217] In one embodiment, the bond between CRM and the compstatin analog includes a carbamate. In one embodiment, the compstatin analog is directly bonded to CRM via a carbamate. In one embodiment, the bond between CRM and the compstatin analog does not include an ester. In one embodiment, the bond between CRM and the compstatin analog includes a carbamate but does not include an ester. In one embodiment, the bond between CRM and the compstatin analog includes a carbamate but does not include a bond that is more susceptible to hydrolysis than the carbamate in an aqueous medium. In one embodiment, CRM includes or is derived from a PEG moiety.

[0218] In one embodiment, the bond between CRM and the compstatin analog includes an amide. In one embodiment, the compstatin analog is directly bonded to CRM via an amide. In one embodiment, the bond between CRM and the compstatin analog includes an amide but does not include an ester. In one embodiment, the bond between CRM and the compstatin analog includes an amide but does not include a bond that is more susceptible to hydrolysis than an amide in an aqueous medium. In one embodiment, CRM includes or is derived from a PEG moiety.

[0219] In one embodiment, one or more compstatin analogs, which are polyfunctionalized compounds (e.g., difunctionalized, trifunctionalized, or more broadly functionalized compounds), are bound to CRM via a bond containing a carbamate. In one embodiment, one or more compstatin analogs, which are polyfunctionalized compounds (e.g., difunctionalized, trifunctionalized, or more broadly functionalized compounds), are bound to CRM via an ester-free bond. In one embodiment, one or more compstatin analogs, which are polyfunctionalized compounds (e.g., difunctionalized, trifunctionalized, or more broadly functionalized compounds), are bound to CRM via a bond containing a carbamate and not a bond that is more susceptible to hydrolysis than the carbamate in an aqueous medium. In one embodiment, each compstatin analog of a polyfunctionalized compound (e.g., difunctionalized, trifunctionalized, or more broadly functionalized compound) is directly bound to CRM via a carbamate.

[0220] In one embodiment, the CRM includes or comprises a PEG moiety. In one embodiment, one or more compstatin analogs that are polyfunctionalized compounds (e.g., difunctionalized, trifunctionalized, or more broadly functionalized compounds) are bonded to the CRM by an amide-containing bond. In one embodiment, one or more compstatin analogs that are polyfunctionalized compounds (e.g., difunctionalized, trifunctionalized, or more broadly functionalized compounds) are bonded to the CRM by an amide-containing, ester-free bond. In one embodiment, one or more compstatin analogs that are polyfunctionalized compounds (e.g., difunctionalized, trifunctionalized, or more broadly functionalized compounds) are bonded to the CRM by an amide-containing, ester-free bond. In one embodiment, each compstatin analog of the polyfunctionalized compound (e.g., difunctionalized, trifunctionalized, or more broadly functionalized compounds) is directly bonded to the CRM via an amide. In one embodiment, the CRM includes or comprises a PEG moiety.

[0221] In one embodiment, the present invention provides compstatin analogs conjugated with a polymer other than PEG. In one embodiment, the polymer is polyoxazoline (POZ). Examples of monofunctional and polyfunctionalized polyoxazoline derivatives for direct or linker-mediated conjugation are given below. ZT-[N(COR x )CH2CH2] n -TR 1 ; R 1 -{[N(CO-TZ)CH2CH2] m -[N(COR x )CH2CH2] n} a -TR 1 ; R 1 -{[N(CO-TZ 1 )CH2CH2] p -[N(COR x )CH2CH2] n -[N(CO-TZ 2 )CH2CH2] m} a -TR 1 ; R 1 -{[N(CO-TZ 1 )CH2CH2] p -[N(COR x )CH2CH2] n -[N(CO-TZ 2 )CH2CH2] m} a -TZ; R 1 -[N(COR x )CH2CH2] n -TB(-R 1 )(-TZ)-T-[N(COR x )CH2CH2] m -R 1 [During the ceremony Z, Z 1 and Z 2 Each of these is a reactive functional group defined above and described here by a group or subgroup; T,R x and R 1 Each of these is defined above and described here as a group or subgroup; Each of m, n, and p is an integer between 0 and 1000, but the sum of m, n, and p in each expression is not 0; a can be "ran" indicating a random copolymer or "block" indicating a block copolymer; B is a branched portion that is linked to the rest of the polymer, either by linkers or without linkers.

[0222] Other examples of functionalized polyoxazoline derivatives for binding include, but are not limited to, those described in PCT patent application publication numbers WO / 2010 / 006282, WO / 2009 / 089542, WO / 2009 / 043027 and WO / 2008 / 106186, the contents of which are incorporated herein by reference.

[0223] An example of a compstatin analog conjugate with a polyoxazoline polymer is shown below. [ka] [ka] [ka] [ka] [In the formula, each variable group is defined above and is described here as a group or subgroup.]

[0224] In one embodiment, the present invention provides polymer-conjugated compstatin analogs, wherein a compstatin analog is bonded to the polymer via one or more linkers. In one embodiment, the polymer is a PEG-containing compound and group described above, and in group or subgroup. In one embodiment, the present invention provides compstatin analog conjugates of the PEG-containing compounds and group of compounds described herein, wherein a compstatin analog is bonded to a PEG-containing portion via one or more linkers. Monofunctional and polyfunctional PEGs containing one or more reactive functional groups for bonding include, but are not limited to, those defined above, and described herein in group or subgroup, and those of formulas A, I, Ia, II, III, IIIa, B, IV, IVa, V, Va, C, D, E, F, G, H, VI, VII, VIII, IX, X, XI, XII, XIII, XIV, XV, or XVI.

[0225] Suitable linkers for linking compstatin analogs to polymer moieties such as PEG or polyoxazoline are broadly described above and here in groups and subgroups. In some embodiments, the linker has multiple functional groups, where one functional group is linked to the compstatin analog and the others to the polymer moiety. In some embodiments, the linker is a bifunctional compound. In some embodiments, the linker is NH2(CH2CH2O) n It has the structure CH2C(=O)OH (where n is 1 to 1000). In one embodiment, the linker is 8-amino-3,6-dioxaoctanoic acid (AEEAc). In one embodiment, the linker is activated for bonding with the functional group of the polymer moiety or compstatin analog. For example, in one embodiment, the carboxyl group of AEEAc is activated before bonding with the amine group of the side chain of the lysine group.

[0226] In one embodiment, a suitable functional group on the compstatin analog (e.g., an amine group, a hydroxyl group, a thiol group, or a carboxylic acid group) is used for bonding to the polymer moiety directly or via a linker. In one embodiment, the compstatin analog is bonded to the PEG moiety via a linker and an amine group. In one embodiment, the amine group is the α-amino group of an amino acid residue. In one embodiment, the amine group is the amine group of a lysine side chain. In one embodiment, the compstatin analog is bonded to NH2(CH2CH2O) n The PEG moiety is bound via the amino group (ε-amino group) of the lysine side chain through a linker having the structure CH2C(=O)OH (where n is 1 to 1000). In one embodiment, a compstatin analog is bound to the PEG moiety via the amino group of the lysine side chain through an AEEAc linker. In one embodiment, NH2(CH2CH2O) n After binding, the CH2C(=O)OH linker forms a -NH(CH2CH2O) group on the compstatin lysine side chain. n The CH2C(=O)- moiety is inserted. In one embodiment, after binding, the AEEAc linker inserts the -NH(CH2CH2O)2CH2C(=O)- moiety into the compstatin lysine side chain.

[0227] In one embodiment, a compstatin analog is bonded to a polymer moiety via a linker, where the linker comprises an AEEAc moiety and an amino acid residue. In another embodiment, a compstatin analog is bonded to a polymer moiety via a linker, where the linker comprises an AEEAc moiety and a lysine residue. In another embodiment, the polymer is PEG. In another embodiment, the C-terminus of the compstatin analog is bonded to the amino group of AEEAc, and the C-terminus of AEEAc is bonded to the lysine residue. In another embodiment, the C-terminus of the compstatin analog is bonded to the amino group of AEEAc, and the C-terminus of AEEAc is bonded to the α-amino group of the lysine residue. In yet another embodiment, the C-terminus of the compstatin analog is bonded to the amino group of AEEAc, and the C-terminus of AEEAc is bonded to the α-amino group of the lysine residue, with a polymer moiety such as a PEG moiety bonded via the ε-amino group of the lysine residue. In yet another embodiment, the C-terminus of the lysine residue is modified. In yet another embodiment, the C-terminus of the lysine residue is modified by amidation. In yet another embodiment, the N-terminus of the compstatin analog is modified. In one embodiment, the N-terminus of the compstatin analog is acetylated.

[0228] Examples of conjugates containing AEEAc linkers and polymers are listed below, where, [ka] This indicates the bonding site of the amine group on the compstatin analog. [ka] However, it exhibits a compstatin analog bound via its C-terminus, and each of the other variable groups is independently defined above and described here in groups and subgroups. In one embodiment, the amine group is the amino group of the lysine side chain. [ka] [ka]

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[0229] In one embodiment, a compstatin analog can be represented as M-AEEAc-Lys-B2, where B2 is a blocking moiety, e.g., NH2, and M is any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A, provided that the C-terminal amino acid of any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A is attached to AEEAc-Lys-B2 via a peptide bond. The NHS moiety of a monofunctional or polyfunctional (e.g., bifunctional) PEG reacts with the free amine of the lysine side chain to produce a monofunctional (single compstatin analog moiety) or polyfunctional (multiple compstatin analog moiety) long-acting compstatin analog. In various embodiments, any amino acid containing a side chain with a reactive functional group can be used instead of (or in addition to) Lys. Monofunctional or polyfunctional PEGs containing appropriate reactive functional groups can be reacted with such side chains in a manner similar to the reaction of NHS-ester activated PEG with Lys.

[0230] In any of the above formulas and structures, embodiments in which the compstatin analog component includes any of the compstatin analogs described herein, for example, any of the compstatin analogs of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, 41A, are expressly disclosed. For example, but not limited to, a compstatin analog may include the amino acid sequence of SEQ ID NO: 28. An example of a long-acting compstatin analog in which the compstatin analog component includes the amino acid sequence of SEQ ID NO: 28 is shown in Figure 10(C). The PEG portion may have a variety of different molecular weights or average molecular weights in various embodiments, as described herein. For example, as described herein, individual PEG chains in a formulation may have different molecular weights and / or different formulations may have different average molecular weights and / or polydispersity. In one embodiment, the PEG portion of the compound in Figure 10(C) has an average molecular weight of approximately 20-100kD, approximately 20-90kD, approximately 20-80kD, approximately 20-70kD, approximately 20-60kD, approximately 20-50kD, approximately 30-80kD, approximately 30-70kD, approximately 30-60kD, approximately 30-50kD, approximately 30-45kD, approximately 35-50kD, approximately 35-45kD, approximately 36-44kD, approximately 37-43kD, approximately 38-42kD, or approximately 39-41kD. In one embodiment, the PEG portion of the compound in Figure 10(C) has an average molecular weight of approximately 30-50kD, for example, approximately 35-45kD, or approximately 37.5-42.5kD. In one embodiment having an average molecular weight of approximately 40 kD, for example, 37.5-42.5 kD, 38 kD, 39 kD, 40 kD, 41 kD, or 42 kD, the compound may be referred to here as CA28-2TS-BF. In one embodiment of a compound containing a CRM, for example a PEG moiety, having an average molecular weight of approximately 40 kD, for example, 37.5-42.5 kD, 38 kD, 39 kD, 40 kD, 41 kD, or 42 kD, the compound has a terminal phase half-life of at least approximately 5 days, for example, approximately 5-10 days, for example, approximately 5, 6, 7, 8, or 9 days, when administered intravenously or subcutaneously to non-human primates or humans at doses of approximately 1-3 mg / kg, 3-5 mg / kg, or 5-10 mg / kg.

[0231] In one embodiment, the present invention relates to the use of click chemistry in relation to compstatin analogs. "Click chemistry" is well known in the art and is useful in one embodiment of the present invention. Click chemistry, in one embodiment, embodies a variety of cycloaddition reactions between azides and alkynes, enabling many useful applications. Methods for carrying out click chemistry are known in the art and are described in their entirety by reference to Kolb, HC; Sharpless, KB, Drug Disc. Today, 2003, 1128-1137; Moses, JE; Moorhouse, AD; Chem. Soc. Rev., 2007, 1249-1262. Click chemistry is a popular method in bioconjugation due to its high reactivity and selectivity, even in biological media. See Kolb, HC; Finn, MG; Sharpless, KB Angew. Chem. Int. Ed. 2001, 40, 2004-2021; and Wang, Q.; Chan, TR; Hilgraf, R.; Fokin, VV; Sharpless, KB; Finn, MGJ Am. Chem. Soc. 2003, 125, 3192-3193. Furthermore, currently available recombinant technologies and synthetic methods enable the introduction of azide and alkyne-containing non-classical amino acids into peptides, proteins, cells, viruses, bacteria, and other biological materials consisting of or displaying proteins. Link, AJ; Vink, MKS; Tirrell, DAJ Am. Chem. Soc. 2004, 126, 10598-10602; Deiters, A.; Cropp, TA; Mukherji, M.; Chin, JW; Anderson, C.; Schultz, PGJ Am. Chem. Soc. 2003, 125, See 11782-11783.

[0232] The term “click chemical group” as used herein may refer to a reactive functional group that can participate in a click chemical reaction with a suitable second reactive functional group, which is also a click chemical group. The first and second click chemical groups, or substances (e.g., molecules) containing such groups, may be called complementary. The first and second substances, e.g., molecules, containing complementary click chemical groups may be called click chemical partners. A substance or molecule containing a click chemical group may be called “click functionalized.” The bond formed by the reaction of complementary click chemical partners may be called a “click chemical bond.”

[0233] In one embodiment, the present invention provides click-functionalized compstatin analogs for, for example, binding to a complementary moiety on a partner molecule or biomolecule. In one embodiment, the complementary partner molecule or biomolecule is a polymer, peptide, protein, or molecule that functions as a clearance reduction moiety. In one embodiment, the “click-functionalized” moiety is an alkyne or alkyne derivative that can be subjected to a [3+2] cycloaddition reaction with a complementary azide-containing molecule and biomolecule. In another embodiment, the “click-functionalized” functionality is an azide or azide derivative that can be subjected to a [3+2] cycloaddition reaction (i.e., click chemistry) with a complementary alkyne-containing molecule and biomolecule.

[0234] In one embodiment, the click-functionalized compstatin analog has an azide group on one of the side chain groups of the compstatin analog. In another embodiment, the click-functionalized compstatin analog has an azide group on the lysine side chain group.

[0235] In one embodiment, the click-functionalized compstatin analog has an alkyne group on one of the side chain groups of the compstatin analog. In another embodiment, the click-functionalized compstatin analog has an alkyne group on the lysine side chain group.

[0236] In one embodiment, the present invention provides a compstatin conjugate comprising a compstatin analog, a molecule functioning as a clearance reduction moiety, and a triazole linker. In one embodiment, the triazole linker is the result of click-bonding chemistry between the compstatin conjugate and the molecule functioning as a clearance reduction moiety. In one embodiment, the CRM may be any of the CRMs disclosed herein. For example, the CRM may be a PEG, polypeptide, or POZ.

[0237] In one embodiment, the present invention provides a compstatin conjugate comprising a compstatin analog, a PEG moiety, and a triazole linker. In one embodiment, the triazole linker is the result of click-bonding chemistry between the compstatin conjugate and the PEG moiety.

[0238] In one embodiment, the present invention provides a compstatin conjugate comprising a compstatin analog, a polyoxazoline moiety, and a triazole linker. In one embodiment, the triazole linker is the result of click-bonding chemistry between the compstatin conjugate and the polyoxazoline moiety.

[0239] In one embodiment, the click chemistry between the compstatin analog and the other part is catalyzed by a transition metal. Copper-containing molecules that catalyze the “click” reaction include, but are not limited to, copper wire, copper bromide (CuBr), copper chloride (CuCl), copper sulfate (CuSO4), copper sulfate pentahydrate (CuSO4·5H2O), copper acetate (Cu2(AcO4)), copper iodide (CuI), [Cu(MeCN)4](OTf), [Cu(MeCN)4](PF6), colloidal copper sources, and immobilized copper sources. In one embodiment, other metals, such as ruthenium, may be used. Reducing agents and organic and inorganic metal-binding ligands may be used with the metal catalyst and include, but are not limited to, sodium ascorbate, tris(triazolyl)amine ligand, tris(carboxyethyl)phosphine (TCEP), batofenanthroline sulfonic acid ligand, and benzimidazole-based ligands.

[0240] In one embodiment, a compstatin analog is bonded to another moiety using metal-free click chemistry (also known as copper-free click chemistry) to obtain a metal-free composition or conjugate. In contrast to standard click chemistry, also known as copper-coated click chemistry (CuACC), metal-free click chemistry occurs between a strained, cyclic alkyne or alkyne precursor, such as oxanorbornadiene, and an azide group. As the name suggests, no metal catalyst is required for the reaction to take place. Examples of such chemistry include reactions involving cyclooctine derivatives (Codelli, et. al. J. Am. Chem. Soc., 2008, 130, 11486-11493; Jewett, et. al. J. Am. Chem. Soc., 2010, 132, 3688-3690; Ning, et. al. Angew. Chem. Int. Ed., 2008, 47, 2253-2255), difluorooxanorbornene derivatives (van Berkel, et. al. ChemBioChem, 2007, 8, 1504-1508), or nitrile oxide derivatives (Lutz, et. al. Macromolecules, 2009, 42, 5411-5413). In some embodiments, metal-free click reactions are metal-free [3+2]cycloaddition reactions, Diels-Alder reactions, or thiol-alkene radical addition reactions. Examples of click reactions and click groups are described, for example, in Joerg Lahann, Click Chemistry for Biotechnology and Materials Science, 2009, John Wiley & Sons Ltd, ISBN 978-0-470-69970-6; and Becer, Hoogenboom, and Schubert, Click Chemistry beyond Metal-Catalyzed Cycloaddition, Angewandte Chemie International Edition (2009) 48: 4900 - 4908. In some embodiments, the click group includes diarylcyclooctyne.

[0241] An example of metal-free click chemistry is shown in the scheme below. [ka]

[0242] Certain metal-free click moieties are known from the literature. An example is 4-dibenzocyclooctinol (DIBO). [ka] (From Ning et al; Angew Chem Int Ed, 2008, 47, 2253); Difluorocyclooctin compounds (DIFO or DFO) [ka] (From Codelli, et. al.; J. Am. Chem. Soc. 2008, 130, 11486-11493); biarylazacyclooctinone (BARAC) [ka] (From Jewett et. al.; J. Am. Chem. Soc. 2010, 132, 3688); or bicyclononine (BCN) [ka] (Dommerholt, et. al.; Angew Chem Int Ed, 2010, 49, 9422-9425) or dibenzylcyclooctin (DBCO) [ka] Includes.

[0243] The reaction scheme, including the reaction between DBCO and azide, is shown below. [ka]

[0244] In the above scheme, in various embodiments, A may include or be derived from a compstatin analog moiety, and B may include or be derived from a CRM, such as a polymer or polypeptide like PEG or POZ, or B may include or be derived from a compstatin analog moiety, and A may include or be derived from a CRM, such as a polymer or polypeptide like PEG or POZ.

[0245] In one embodiment, the “metal-free click-functionalized” portion is acetylene or an acetylene derivative that can be subjected to a [3+2] cycloaddition reaction with complementary azide-containing molecules and biomolecules without the use of a metal catalyst.

[0246] In one embodiment, the R and R' groups of the metal-free click chemical reagent may be a compstatin analog or any of the molecules described herein to which the compstatin analog can be bound. In one embodiment, such a compstatin analog has a click-functionalized moiety on the lysine side chain. In one embodiment, such a compstatin analog is bound to the click-functionalized moiety via a linker. In one embodiment, such a compstatin analog is bound to the click-functionalized moiety via AEEAc.

[0247] In one embodiment, the click chemical reagent includes DBCO. Examples of the reagent and its use are shown below. [ka] DBCO-acid. In one embodiment, DBCO-acid may be used for reaction with an amine-containing moiety.

[0248] [ka] DBCO-NHS ester (above) or DBCO-sulfo-NHS ester (below), DBCO functionality can be used to incorporate amine-containing molecules such as compstatin analogs or polypeptides that contain lysine residues. [ka]

[0249] [ka] DBCO-PEG4-NHS ester. In one embodiment, such a reagent is useful for incorporating the DBCO moiety by reaction with available amine functionalities. In one embodiment, the presence of a PEG chain as a hydrophilic spacer may be useful, for example, to increase solubility or provide flexibility.

[0250] [ka] DBCO-amine. In one embodiment, the click chemical reagent comprises carbonyl / carboxyl-reactive dibenzylcyclooctin that can react with acids, active esters and / or aldehydes.

[0251] In one embodiment, the click chemical reaction includes the following cyclooctane. [ka]

[0252] In one embodiment, the click chemical reaction involves the reaction of nitrone and cyclooctin (see, e.g., Ning, Xinghai; Temming, Rinske P.; Dommerholt, Jan; Guo, Jun; Ania, Daniel B.; Debets, Marjoke F.; Wolfert, Margreet A.; Boons, Geert-Jan et al. (2010). “Protein Modification by Strain-Promoted Alkyne-Nitrone Cycloaddition”. Angewandte Chemie International Edition 49 (17): 3065), aldehydes and ketones, oxime / hydrazone formation from tetrazine ligation (see, e.g., Blackman, Melissa L.; Royzen, Maksim; Fox, Joseph M. (2008). “The Tetrazine Ligation: Fast Bioconjugation based on Inverse-electron-demand Diels-Alder Reactivity”. Journal of the American Chemical Society 130 (41): See 13518-9), tetrazole ligation, isonitrile-based click reactions (e.g., Stackmann, Henning; Neves, Andre A.; Stairs, Shaun; Brindle, Kevin M.; Leeper, Finian J. (2011). “Exploring isonitrile-based click chemistry for ligation with biomolecules”. See Organic & Biomolecular Chemistry 9 (21): 7303), and quadricyclane ligation (e.g., Sletten, Ellen M.; Bertozzi, Carolyn R. (2011). “A Bioorthogonal Quadricyclane Ligation”.(See Journal of the American Chemical Society 133 (44): 17570-3). In one embodiment, the click reaction is a Staudinger ligation (phosphine-azide).

[0253] Any compstatin analog can be modified to incorporate a click group in various embodiments. For example, a compstatin analog containing any of the sequences of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A can be modified in this way. In some embodiments, any such sequence further includes a lysine residue or an AEEAc-Lys moiety, for example, at the C-terminus. In some embodiments, the click group is inserted after peptide synthesis. For example, the Lys side chain may be reacted with azidoacetic acid to insert an azide moiety as a click group. In some embodiments, the click group is inserted after cyclization, and in some embodiments, after the addition of N-terminal and / or C-terminal blockade moieties. In some embodiments, the click group is inserted during peptide synthesis. For example, an amino acid containing a side chain with a click group can be used in the synthesis of a compstatin analog. A variety of such amino acids are commercially available from many sources, for example, AAPPTec (Louisville, KY) and Jena Bioscience GmbH (Jena, Germany). In one embodiment, a method for producing click-functionalized compstatin analogs is provided hereby.

[0254] In one embodiment, a composition comprising a compstatin analog and a click chemical reagent is provided. The click chemical reagent may be any molecule that can react with the amino acid side chain or terminal of the compound comprising the compstatin analog to introduce a click chemical group, e.g., any click chemical group known in the art. In one embodiment, the composition may be incubated under suitable conditions for functionalizing the compstatin analog with click chemical functionality (which may include providing a suitable catalyst and light (e.g., UV)). In one embodiment, the present invention provides a compstatin analog comprising any click chemical group, including but not limited to those described herein. In one embodiment, a method for producing a long-acting compstatin analog is provided. In one embodiment, the method comprises mixing a compstatin analog comprising a first click chemical group with a CRM comprising a complementary click chemical group under conditions suitable for the click chemical reaction to occur. The method may further include purifying the resulting conjugate. In one embodiment, the purification comprises removing at least some unreacted components, e.g., with a suitable scavenger.

[0255] In one embodiment, a click reaction is used to link two or more CRMs, with at least two of them having a compstatin analog moiety. The compstatin analog moiety may be the same or different in various embodiments. The compstatin analog moiety may or may not be bound to the CRM via the click reaction. For example, in one embodiment, a first heterobifunctional PEG containing a first click chemical group at the first end and an NHS ester at the second end is coupled to the compstatin analog moiety via the NHS ester. In another reaction, a second heterobifunctional PEG containing a second click chemical group at the first end and an NHS ester at the second end is coupled to the compstatin analog moiety via the NHS ester. The two resulting compounds are then reacted via a click reaction to form a large molecule containing two compstatin analog moieties. Although PEG is described as an example of a CRM, this method should be interpreted as being able to use any CRM. For example, in one embodiment, a polypeptide, e.g., HSA or a part thereof, or albumin or an albumin-binding peptide, or an antibody or a part thereof, may be used as a CRM. In one embodiment, POZ can be used in this manner.

[0256] Compstatin analogs containing click groups can have diverse applications. In one embodiment, a compstatin analog containing a primary click group is reacted with any substance containing a complementary click group. Substances containing complementary click groups may include, for example, labels (e.g., fluorophores, fluorescent proteins, radioisotopes), affinity reagents, antibodies, targeting moieties, metals, particles, etc. In one embodiment, a click group is used to bond a compstatin analog moiety to a surface, which contains or is functionalized to contain a complementary click group. In one embodiment, the surface is a sensor, e.g., a surface or sensor for capturing / detecting C3. In one embodiment, the surface forms part of a medical device, tubes, membranes, reservoirs, implants, or other substance that may come into contact with blood (e.g., outside the body) or may be temporarily or permanently implanted in the body of a subject (e.g., an artificial organ or drug delivery device). In one embodiment, the surface is functionalized with a compstatin analog to reduce complement activation to it. In one embodiment, the device or tubing is used for blood circulation, for example, in dialysis or during surgery. In another embodiment, the device is a hemodialyzer or an extracorporeal circulation support unit. Such a Compstatin analog-functionalized device and a method for manufacturing the same are provided herein.

[0257] In one embodiment of the present invention, the compstatin analog comprises both a cell-reactive functional group and a CRM. In one embodiment, the present invention comprises a cell-reactive functional group or moiety having a molecular weight of at least 500 daltons, for example, at least 1,500 daltons to a maximum of about 100,000 daltons (e.g., an average molecular weight of about 20,000; 30,000; 40,000; 50,000; 60,000; 70,000; 80,000; 90,000; or 100,000 daltons) (CH2CH2O). n The present invention provides a variant of any of the above cell-reactive compstatin analog molecules that replaces a portion (e.g., any of the PEGs described herein) or another polymer (e.g., POZ, polypeptide). In one embodiment, the compound or (CH2CH2O)n The average molecular weight of the moiety (or other polymer, e.g., POZ or polypeptide) is at least 20,000 daltons and up to approximately 100,000; 120,000; 140,000; 160,000; 180,000; or 200,000 daltons. Therefore, the teachings herein regarding the binding of cell-reactive compstatin analogs, e.g., the compstatin analog moiety used and the binding of the compstatin analog moiety to the cell-reactive moiety, can be provided for long-acting compstatin analogs, which may have any of the structures shown by ALM as described above, where A is a clearance-reducing moiety (e.g., any of the clearance-reducing moieties described herein), and further, where L (or L P1 , L P2 , or L P3There may be one or more (e.g., 3, 4, 5, 6, 7, 8) compstatin analog moieties M that bind to A via a binding site shown as ). The compstatin analog moieties may include a peptide or a variant thereof (e.g., any of the variants described herein) whose sequence includes any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A, which may be extended by one or more amino acids at the N-terminus, C-terminus, or both ends, wherein at least one amino acid has a side chain containing a reactive functional group such as a primary or secondary (e.g., Lys), a sulfhydryl group, a carboxyl group (which may exist as a carboxylic acid group), a guanidino group, a phenol group, an indole ring, a thioether, or an imidazole ring, which facilitates the conjugation of the CRM-containing moiety to the compstatin analog (it is understood that after conjugation, such reactive functional groups react to form a bond). Furthermore, a compstatin analog moiety or variant containing any of SEQ ID NOs. 3-36, 37, 37A, 38A, 39A, 40A, or 41A is extended by one or more amino acids at its N-terminus, C-terminus, or both ends, where at least one of the amino acids has a side chain containing a reactive functional group, such extension of one or more amino acids is derived from the cyclic portion of the compstatin analog moiety, for example, a substituted or unsubstituted, saturated or unsaturated alkyl chain, an oligo(ethylene glycol) chain, and / or L(L) P1 , L P2 , or L P3 It is also understood that they may be separated by flexible or rigid spacer portions, including any of the other parts indicated as ).

[0258] Examples of long-acting compstatin analogs where n is a value sufficient to obtain an average molecular weight of approximately 500;1,000;1,500;2,000;5,000;10,000;20,000;30,000;40,000;50,000;60,000;70,000;80,000;90,000;~100,000 daltons are listed below. In one embodiment, n is a value sufficient to provide an average molecular weight of approximately 20,000 daltons, up to approximately 100,000;120,000;140,000;160,000;180,000; or 200,000 daltons.

[0259] (CH2CH2O) n C(=O)-Ile-Cys-Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys-Thr-NH2)(Sequence ID 58) Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH-CH2CH2OCH2CH2OCH2-C(=O)-Lys-C(=O)-(CH2CH2O) n -NH2 (SEQ ID NO: 59) Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-Lys-C(=O)-(CH2CH2O) n -NH2 (Sequence ID 60). Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-(Gly)5-Lys-C(=O)-(CH2CH2O) n -NH2 (Sequence ID 61) Ac-(CH2CH2O) n C(=O)Lys-(Gly)5-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH2) (Sequence ID 62) Ac-(CH2CH2O) n C(=O)Lys-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH 2) (Sequence No. 63)

[0260] In sequence number 58, (CH2CH2O) n It is bonded to the N-terminal amino acid via an amide bond. In sequence numbers 59-63, (CH2CH2O) n The portion is bonded to the Lys side chain via an amide bond. Therefore, as described above, the C-terminal NH2 in SEQ ID NOs. 59, 60, and 61 is understood to represent the C-terminal amidation of the peptide, and in SEQ ID NOs. 62 and 63, the N-terminal Ac is understood to represent the N-terminal acetylation of the peptide. In addition, (CH2CH2O) n The free ends of the part are usually underlined O, and the O represents the O atom of the terminal (CH2CH2O) group. O It will be understood by those skilled in the art that it ends in R). (OR (underlined O)) is often hydroxyl O H) group or methoxy(- O While the CH3 group is a specific part, other groups (for example, other alkoxy groups) can also be used. Therefore, for example, SEQ ID NO: 59 is an Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH-CH2CH2OCH2CH2OCH2-C(=O)-Lys-(C(=O)-(CH2CH2O) nIt can be represented as -R)-NH2(SEQ ID NO: 64) (wherein R is, for example, H or CH3 in the case of linear PEG). In the case of bifunctional, branched or star-shaped PEG, R represents the rest of the molecule. Furthermore, it is understood that the portion containing the reactive functional group may be modified as described herein (for example, according to any of the formulas described herein). For example, a long-acting compstatin analog in which the portion containing the reactive functional group contains an ester and / or alkyl chain and has the same peptide sequence as SEQ ID NO: 64 can be represented as follows: Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH-CH2CH2OCH2CH2OCH2-C(=O)-Lys-(C(=O)-(CH2) m -(CH2CH2O) n -R)-NH2(sequence number 65); Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH-CH2CH2OCH2CH2OCH2-C(=O)-Lys-(C(=O)-(CH2) m- C(=O)-(CH2CH2O) n -R)-NH2 (Sequence ID 66) Ac-Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr-NH-CH2CH2OCH2CH2OCH2-C(=O)-Lys-(C(=O)-(CH2) m- C(=O)-(CH2)j (CH2CH2O) n -R)-NH2 (Sequence ID 67)

[0261] In sequence numbers 65-67, m can range from 1 to approximately 2, 3, 4, 5, 6, 7, 8, 10, 15, 20, or 30 in various embodiments. In sequence number 67, j can range from 1 to approximately 2, 3, 4, 5, 6, 7, 8, 10, 15, 20, or 30 in various embodiments. Also, as described here, in various embodiments, Lys-(C(=O)- and (CH2CH2O) n It is also understood that other parts, such as amides, aromatic rings (e.g., substituted or unsubstituted phenyl), or substituted or unsubstituted cycloalkyl structures, may be incorporated between -R.

[0262] The present invention provides variants of sequence numbers 58-67, which are -Ile-Cys * -Val-(1Me)Trp-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * -Thr- is replaced by an amino acid sequence containing any other compstatin analog, such as any of the amino acid sequences of SEQ ID NOs: 3-27 or 29-36, 37, 37A, 38A, 39A, 40A, or 41A, provided that the blocking moiety present at the N-terminus and / or C-terminus of the compstatin analog is absent, replaced by a linker (which may contain the blocking moiety), or bound to a different N-terminal or C-terminal amino acid present within the corresponding variant.

[0263] Any compstatin analog, for example, any compound comprising any of SEQ ID NOs: 3-37, 37A, 38A, 39A, 40A, or 41A, may, in various embodiments, directly or indirectly bind to any moiety containing a reactive functional group, for example, any compound of formulas I-XVI or A-H, via or near its N-terminus or C-terminus (for example, via the side chain of an amino acid near its N-terminus or C-terminus).

[0264] In one embodiment, CRM comprises a polypeptide or fragment thereof found in human serum, or a variant substantially similar to the above polypeptide or fragment thereof. In one embodiment, the polypeptide, fragment, or variant has a molecular weight of 5 to 150 kD, for example, at least 5 kD, 10 kD, 20 kD, 30 kD, 40 kD, 50 kD, 60 kD, 70 kD, 80 kD, 90 kD, 100 kD or more, for example, 100 to 120 kD or 120 to 150 kD. In one embodiment, the production of a long-acting compstatin analog comprises reacting a compstatin analog containing a reactive functional group with one or more amino acid side chains of a polypeptide, wherein the side chains contain compatible functional groups. In one embodiment, the production of a long-acting compstatin analog comprises reacting a compstatin analog containing a reactive functional group with the N-terminal amine and / or C-terminal carboxyl group of a polypeptide. In one embodiment, the production of a long-acting compstatin analog comprises reacting a compstatin analog containing an amine-reactive functional group with an N-terminal amine of an amino acid and / or polypeptide having a side chain containing a primary amine (e.g., lysine). In another embodiment, the production of a long-acting compstatin analog comprises reacting a compstatin analog containing a carboxyl-reactive functional group with a C-terminal carboxyl group of a polypeptide. In yet another embodiment, the compstatin analog moiety is attached to each end of a polypeptide, and optionally to the side chains of one or more internal amino acids. In yet another embodiment, the production of a long-acting compstatin analog comprises reacting a compstatin analog containing a sulfidyl-reactive functional group with one or more sulfidyl groups of a polypeptide.

[0265] In one embodiment, at least one reactive functional group is introduced into the polypeptide. For example, in one embodiment, at least one side chain of the polypeptide is modified to convert the first reactive functional group to a different reactive functional group before reacting with a compstatin analog. In one embodiment, a thiol is introduced. Several methods can be used to introduce a thiol into a biomolecule, including the reduction of endogenous disulfides and the conversion of amine groups, aldehyde groups, or carboxylic acid groups to thiol groups. Disulfide crosslinks of cystine in proteins can be reduced to cysteine ​​residues with dithiothreitol (DTT), tris-(2-carboxyethyl)phosphine (TCEP), or tris-(2-cyanoethyl)phosphine. Amines can be indirectly thiolated by reacting with succinimimidyl 3-(2-pyridyldithio)propionate (SPDP) and then reducing the 3-(2-pyridyldithio)propionyl conjugate with DTT or TCEP. The amine can be indirectly thiolated by reacting it with succinimidylacetylthioacetate and then removing the acetyl group with 50 mM hydroxylamine or hydrazine at a nearly neutral pH. The amine can be directly thiolated by reacting it with 2-iminothiolane and introducing a free thiol while maintaining the overall charge of the molecule. Tryptophan residues in thiol-free proteins can be oxidized to mercaptotryptophan residues, which can then be modified with iodoacetamide or maleimide. Polypeptides containing one or more thiols and Ac-Ile-Cys * -Val-Trp(1-Me)-Gln-Asp-Trp-Gly-Ala-His-Arg-Cys * Long-acting compstatin analogs may be produced by reacting them with compstatin analogs containing a maleimide group, such as -Thr-AEEAc-Lys-(C(=O)-(CH2)5-Mal)-NH2 (SEQ ID NO: 68).

[0266] In one embodiment, the polypeptide is produced by recombinant manufacturing. In one embodiment, the polypeptide is produced at least partially by recombinant manufacturing (e.g., in bacteria, or in the cells of a eukaryotic host such as a fungus, insect, plant, or vertebrate) and / or at least partially by chemical synthesis. In one embodiment, the polypeptide is a purified product. For example, in one embodiment, the polypeptide is purified from a host cell lysate or from a culture medium secreted by a host cell. In one embodiment, the polypeptide is glycosylated. In one embodiment, the polypeptide is not glycosylated. In one embodiment, the polypeptide is human serum albumin (HSA). In one embodiment, a variant substantially similar to the polypeptide is similar to the polypeptide to such an extent that it is not recognized as a foreign substance by the normal immune system of a subject, e.g., a human subject. In one embodiment, a variant sequence change that is substantially similar to the original polypeptide is selected so as not to result in an MHC class I epitope. Various methods known in the art can be used to predict whether a sequence contains an MHC class I epitope.

[0267] In some embodiments, one or more amino acids in a polypeptide or linker or composition may be selected to be hydrophobic or hydrophilic, or selected to confer high hydrophilicity or, in some embodiments, high hydrophobicity to a compound containing them. As is well known in the art, the terms “hydrophilic” and “hydrophobic” are used to describe the degree of affinity a substance has for water. In some embodiments, hydrophilic substances have a strong affinity for water and tend to dissolve in water, mix with water, or become wet with water, while hydrophobic substances have substantially no affinity for water, tend to repel and not absorb water, and tend not to dissolve in water, not mix with water, or become wet with water. Amino acids are classified on the basis of hydrophobicity, as is well known in the art. Examples of “hydrophilic amino acids” are arginine, lysine, threonine, alanine, asparagine, glutamine, aspartic acid, glutamic acid, serine, and glycine. Examples of “hydrophobic amino acids” are tryptophan, tyrosine, phenylalanine, methionine, leucine, isoleucine, and valine. In one embodiment, an analog of a standard amino acid is used, wherein the analog has increased or decreased hydrophilicity or hydrophobicity compared to the amino acid from which the analog is derived.

[0268] The present invention further provides a polymer, e.g., concatemer, containing two or more (e.g., 2 to 10) CRM-containing compstatin analogs, where the average molecular weight of the resulting molecule (or its CRM component) is 20,000; 30,000; 40,000; 50,000; 60,000; 70,000; 80,000; 90,000; ~100,000 daltons. In one embodiment, the average molecular weight of the resulting molecule (or its CRM component) is at least 20,000 daltons and at most about 100,000; 120,000; 140,000; 160,000; 180,000 or 200,000 daltons. In one embodiment, any of the above binding moieties can be used to bind the CRM-containing compstatin analog. Compositions, methods, and synthetic intermediates for producing long-acting compstatin analogs are one aspect of the present invention.

[0269] In one embodiment, the total molecular weight of the long-acting compstatin analog containing the compstatin analog moiety does not exceed 50 kD. For example, in the case of LACA containing 40 kD PEG, in one embodiment, the molecular weight contributed by the remaining portion of the compound containing the compstatin analog moiety does not exceed 10 kD, e.g., 1.5 kD to 5.0 kD or 5.0 kD to 10 kD. In one embodiment, the total molecular weight of LACA containing the compstatin analog moiety is 45 kD to 50 kD. In one embodiment, the total molecular weight of LACA containing the compstatin analog moiety is 40 kD to 45 kD, 15 kD to 40 kD, e.g., 15 kD to 25 kD, 25 kD to 35 kD, 35 kD to 40 kD. Therefore, when the present invention refers to a compstatin analog comprising a polymer or CRM having a specific molecular weight or a molecular weight within a specific range, in some embodiments the total molecular weight of the compstatin analog may be, for example, 1.5 kD to 5 kD greater than the molecular weight of the polymer or CRM, or in some embodiments, 5 kD to 10 kD greater than the molecular weight of the polymer. The molecular weight of a compound, for example a compound comprising a polymer, may refer to the average molecular weight of the molecules of such a compound in the composition.

[0270] A wide range of methods and assays are known in the art that are useful for the detection of polymers, such as PEG, POZ, and / or polypeptides, and for the measurement of the physical and / or structural properties of polymers, such as PEG, POZ, and / or polypeptides, and may be used for the detection of compstatin analogs, such as cellular reactivity, long-acting, targeted compstatin analogs, or compstatin analog moieties. For example, methods and assays are available that are useful for measuring properties such as aggregation, solubility, size, structure, melting properties, purity, presence of degradation products or contaminants, water content, and hydrodynamic radius. Such methods include, for example, centrifugal analysis, various types of chromatography such as liquid chromatography (e.g., HPLC-ion exchange, HPLC-molecular sieving, HPLC-reverse phase), light scattering, capillary electrophoresis, circular dichroism, isothermal calorimetry, differential scanning calorimetry, fluorescence, infrared (IR), nuclear magnetic resonance (NMR), Raman spectroscopy, refractive index measurement, UV / visible spectroscopy, mass spectrometry, and immunological methods. It is understood that methods can be combined. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog (or a composition comprising a cell-reactive, long-acting, or targeted compstatin analog) is evaluated using any of the aforementioned methods and possesses one or more of the properties described herein. In one embodiment, a method useful for the detection and / or quantification of long-acting compstatin analogs is described herein.

[0271] VI. Targeted Compstatin Analogues The present invention provides and / or utilizes a targeted compstatin analog comprising a targeting moiety and a compstatin analog moiety, wherein the targeting moiety non-covalently binds to a target molecule. In some aspects, the present invention provides a targeted compstatin analog similar to the cell-reactive compstatin analog described in Section VI, wherein the compound comprises a targeting moiety in addition to, or instead of, the cell-reactive moiety. The targeting moiety may include, for example, antibodies, polypeptides, peptides, nucleic acids (e.g., aptamers), carbohydrates, small molecules, or supramolecular complexes that specifically bind to a target molecule. In one embodiment, the affinity of the targeting moiety (measured by the equilibrium dissociation constant Kd) to the target molecule (measured by the equilibrium dissociation constant Kd) is 10 under test conditions, for example, under physiological conditions. -3 M or less, for example, 10 -4 M or less, for example, 10 -5 M or less, for example, 10 -6 M or less, 10 -7 M or less, 10 -8 M or less or 10 -9 It is M or less.

[0272] In embodiments of the present invention where the targeted portion is an antibody, the antibody may be any immunoglobulin or its derivative that retains binding ability, or any protein having a binding domain homologous or nearly homologous to an immunoglobulin-binding domain. Such proteins may be derived from natural sources, or may be synthesized in part or in whole (e.g., using recombinant DNA technology, chemical synthesis, etc.). The antibody may be from any species, e.g., human, rodent, rabbit, goat, chicken, etc. The antibody may be a member of any immunoglobulin class, including all human classes: IgG, IgM, IgA, IgD, and IgE. In various embodiments of the present invention, the antibody may be an antibody fragment such as Fab', F(ab')2, scFv (single-chain variable region), or other fragments retaining an antigen-binding site, or a recombinant scFv fragment including a recombinantly produced fragment. See, for example, Allen, T., Nature Reviews Cancer, Vol.2, 750-765, 2002 and its references. Monovalent, bivalent, or polyvalent antibodies can be used. The antibody may be a chimeric antibody, for example, in which a variable domain derived from a rodent and a constant domain derived from a human are fused, thereby retaining the specificity of a rodent antibody. In one embodiment, a human antibody or a portion thereof is produced in a rodent, for example, in which a human immunoglobulin gene is incorporated into the genome, using a display technique such as phage display. In another embodiment, a humanized antibody is produced by transplanting one or more complementarity-determining regions derived from a non-human species (e.g., mouse) into a human antibody sequence. The antibody may be partially or entirely humanized. For example, see Almagro JC, Fransson J., Humanization of antibodies. Front Biosci. 13: 1619-33 (2008) for an overview of various methods for obtaining humanized antibodies that can be used to obtain a targeting portion useful for the present invention. Monoclonal antibodies are generally preferred for the purposes of the present invention, but the antibody may be polyclonal or monoclonal.In one embodiment of the present invention, an F(ab')2 or F(ab') fragment is used, and in another embodiment, an antibody containing an Fc domain is used. Methods for producing antibodies that specifically bind to virtually any molecule of interest are known in the art. For example, monoclonal or polyclonal antibodies can be produced from natural sources, for example, by purification from the blood or ascites fluid of an animal producing the antibody (for example, after immunosensitization with the molecule or its antigenic fragment), or by recombinant cell culture. Methods for producing antibody fragments are known in the art, for example, by digestion, disulfide reduction, or synthesis.

[0273] In various embodiments of the present invention, the targeted moiety may be any molecule that specifically binds to a target molecule by a mechanism other than antigen-antibody interaction. Such a targeted moiety is called a “ligand.” For example, in various embodiments of the present invention, the ligand may be a polypeptide, peptide, nucleic acid (e.g., DNA or RNA), carbohydrate, lipid or phospholipid, or small molecule. In some embodiments, the small molecule is an organic compound, whether natural or artificially produced, that has a relatively low molecular weight, is not a protein, polypeptide, nucleic acid, or lipid, typically has a molecular weight of less than about 1500 g / mol, and typically has multiple carbon-carbon bonds. Generally, an aptamer is an oligonucleotide (such as RNA or DNA) that binds to a specific protein, for example, by stabilizing the molecule by increasing its resistance to degradation by nucleases, for example, by modified nucleosides (e.g., bases or sugars other than the five standard bases (A, G, C, T, U) or sugars (ribose and deoxyribose) most commonly found in RNA and DNA) or modified nucleoside bonds (e.g., non-phosphodiester bonds) in the case of RNA or DNA. In one embodiment, the oligonucleotide has a maximum length of approximately 100 nucleosides, for example, 12 to 100 nucleosides. Aptamers can be obtained using an in vitro evolution method called SELEX, and methods for obtaining aptamers specific to a target protein are known in the art. See, for example, Brody EN, Gold L. J Biotechnol. 2000 March; 74(1): 5-13. In one embodiment, peptide nucleic acids or located nucleic acids are used.

[0274] In one embodiment of the present invention, the targeting moiety includes a peptide. In one embodiment, a display technique such as a phage display, ribosome display, or yeast display is used to identify the peptide that binds to the target molecule of interest.

[0275] Small molecules can be used as ligands. Methods for identifying such ligands are well known in the art. For example, in vitro screening of small molecule libraries, including combinatorial libraries, and computer-based screening for identifying low molecular weight organic compounds that bind to the concave surfaces (pockets) of proteins can identify numerous small molecule ligands for target proteins (Huang, Z., Pharm. & Ther. 86: 201-215, 2000).

[0276] In some embodiments of the present invention, the targeted portion is not a protein or molecule that is typically used as a carrier to conjugate with an antigen for the purpose of inducing an antibody. Examples include carrier proteins or molecules such as bovine serum albumin, keyhole limpet hemocyanin, bovine gamma globulin, and diphtheria toxin. In some embodiments of the present invention, the targeted portion is not the Fc portion of an immunoglobulin molecule. In some embodiments, the targeted portion is part of a complex that includes one or more additional portions to which it is covalently or noncovalently bonded.

[0277] In various embodiments of the present invention, the target molecule may be any molecule produced by a cell (including any form expressed on the cell surface or any modified form in which at least part is due to extracellular modification). In some embodiments, the target molecule is an extracellular substance present in or on a tissue. In some embodiments, the target molecule is characterized by a particular disease or physiological condition, or by one or more cell types or tissue types. Often, the target molecule is at least partially present on the cell surface (e.g., a transmembrane protein or membrane-bound protein) so that at least a portion of the molecule is readily bound by an extracellular binding substance such as an antibody. The target molecule may, but does not need to be, cell type specific. For example, cell type specific target molecules are often proteins, peptides, mRNA, lipids, or carbohydrates that are present at higher levels on or within one or more specific types of cells than on or within many other types of cells. In some cases, the cell type specific target molecule is present at detectable levels only within or on the specific type of cell of interest. However, it is understood that a useful cell type specific target molecule does not need to be perfectly specific to the target cell type to be considered cell type specific. In one embodiment, a cell type-specific target molecule of a particular cell type is expressed within that cell type at a level at least three times higher than that of a reference cell population consisting of a mixture of cells from multiple (e.g., 5 to 10 or more) different tissues or organs in roughly equal amounts. In another embodiment, the cell type-specific target molecule is present at a level at least 4 to 5 times, 5 to 10 times, or 10 times or more higher than its average expression level in the reference population. In yet another embodiment, a person skilled in the art can distinguish one or more target cell types from many other types, most types, or all types of cells by detecting or measuring the cell type-specific target molecule.Generally, the presence and / or quantity of most target molecules can be determined using one or more standard techniques, such as Northern blotting, in situ hybridization, RT-PCR, sequencing, immunological methods (immunoblotting, immunodetection (e.g., by immunohistochemistry), or fluorescence detection after staining with fluorescently labeled antibodies (e.g., using FACS)), oligonucleotide microarrays, cDNA microarrays, membrane arrays, protein microarray analysis, and mass spectrometry.

[0278] In one embodiment, the target molecule is a channel, transporter, receptor, or other molecule in which at least a portion is exposed on the cell surface. In another embodiment, the target molecule is an anion transporter or water channel (e.g., an aquaporin protein).

[0279] In one embodiment, the target molecule is a glycophorin (e.g., glycophorin A, B, C, or D) or a protein such as band 3 in which at least a portion is exposed on the surface of the red blood cell.

[0280] In one embodiment, the target molecule is a protein that is at least partially exposed on the surface of endothelial cells. In one embodiment, the target molecule is present on the surface of a normal, healthy vascular system. In one embodiment, the target molecule is present on the surface of activated endothelial cells. In one embodiment, the target molecule is present on the surface of activated endothelial cells but not on the surface of inactivated endothelial cells. In one embodiment, the target molecule is a molecule whose expression or exposure is induced by stimuli such as injury or inflammation. In one embodiment, the target molecule may be recognized as “non-self” by a recipient who has received a graft containing cells expressing the target molecule. In one embodiment, the target molecule is a heterologous carbohydrate antigen against which antibodies are commonly found in humans. In one embodiment, the carbohydrate includes blood group antigens. In one embodiment, the carbohydrate includes heterologous antigens. For example, the alpha-gal epitope (Gal-alpha1-3Gal-beta1-(3)4GlcNAc-R) (see, for example, Macher BA and Galili U. The Galalpha1, 3Galbeta1, 4GlcNAc-R (alpha-Gal) epitope: a carbohydrate of unique evolution and clinical relevance. Biochim Biophys Acta. 1780(2): 75-88 (2008)).

[0281] In one embodiment of the present invention, the compstatin analog comprises both the targeting moiety and the CRM. In one embodiment, a targeted compstatin analog comprises multiple targeting moieties, which may be the same or different. Different targeting moieties may bind to the same target molecule or to different target molecules. The present invention provides a targeted compstatin analog that is polyvalent with respect to the targeting moiety, the compstatin analog, or both.

[0282] In general, the present invention encompasses all methods for producing compounds comprising a compstatin analog moiety and a targeting moiety, and the compounds obtained thereby. In some embodiments, targeted compstatin analogs can be produced using methods generally similar to those generally described in Chapter VI, but here the targeting moiety is used instead of, or in addition to, the cell-reactive moiety. In some embodiments, a targeted compstatin analog comprising a peptide as the targeting moiety is synthesized as a polypeptide chain comprising a compstatin analog moiety and a peptide targeting moiety. Optionally, the polypeptide chain contains one or more spacer peptides between the compstatin analog moiety and the targeting moiety.

[0283] In one embodiment, a targeted compstatin analog has at least about 10%, 20%, or 30% of the activity of a corresponding compstatin analog having the same amino acid sequence (and, where applicable, one or more blockage moieties) but without the targeted moiety, e.g., 30-40%, 30-50%, 30-60%, 30-70%, 30-80%, 30-90%, or more molar activity. In one embodiment, the targeted compstatin analog comprises multiple compstatin analog moieties, the molar activity of the targeted compstatin analog is at least about 10%, 20%, or 30% of the total activity of the compstatin analog moieties, e.g., 30-40%, 30-50%, 30-60%, 30-70%, 30-80%, 30-90%, or more molar activity of the targeted compstatin analog moieties. Compositions of targeted compstatin analogs, methods for producing them, and synthetic intermediates are aspects of the present invention.

[0284] VII. Use Cell-reactive, long-acting, or targeted compstatin analogs have a wide variety of applications. Specific applications of cell-reactive, long-acting, or targeted compstatin analogs and related embodiments of the present invention are described herein, without limiting the scope of the invention. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is administered to a subject suffering from or at risk of complement-mediated injury to an organ, tissue, or cell. In another embodiment, the cell-reactive compstatin analog is ex vivo into contact with and covalently binds to an organ, tissue, or cell. The organ, tissue, or cell is introduced into the subject and protected from damage that may be induced by the recipient's complement system.

[0285] For the purposes described herein, non-covalently binding compstatin analogs can be used. For example, compstatin analogs modified with a moiety that extends the lifespan of the compound in the body and / or compstatin analogs that include a moiety that targets the compstatin analog to cell types or sites susceptible to complement activation can be used, and the present invention encompasses such uses. In one embodiment, a long-acting compstatin analog is used. In one embodiment, a compstatin analog containing a targeting moiety is used. In one embodiment, a compstatin analog containing both a moiety that extends the lifespan of the compound in the body and a targeting moiety is used. Where cell-reactive compstatin analogs are referred to below, the present invention provides similar compositions and methods relating to targeted compstatin analogs, as well as embodiments (at least in embodiments relating to the administration of compstatin analogs to a target) that use a compstatin analog without a targeting moiety or a cell-reactive moiety, optionally a long-acting compstatin analog, instead of, or in addition to, a cell-reactive compstatin analog.

[0286] Intended uses include: (1) protecting RBCs from complement-mediated injury in patients with disorders such as paroxysmal nocturnal hemoglobinuria or atypical hemolytic uremic syndrome or other disorders characterized by complement-mediated erythrocyte (RBC) lysis; (2) protecting transplanted organs, tissues, and cells from complement-mediated injury; (3) mitigating I / R injury (e.g., in patients in situations where trauma, vascular occlusion, myocardial infarction, or other ischemia / reperfusion (I / R) injury is likely to occur); and (4) protecting various body structures (e.g., the retina) or membranes (e.g., the synovial membrane) that may be exposed to complement components from complement-mediated injury in any of the various different complement-mediated disorders. The beneficial effects obtained by inhibiting complement activation on the surface of cells or other body structures are not limited to those directly resulting from protecting the cells or structures themselves from complement-mediated injury (e.g., prevention of cytolysis). For example, inhibiting complement activation using cell-reactive compstatin analogs may reduce anaphyllotoxin production and the resulting neutrophil influx / activation and other pro-inflammatory events, and / or reduce the release of potentially damaging cellular contents, potentially leading to beneficial effects on distant organ systems or the entire body.

[0287] A. Blood cell protection In one embodiment of the present invention, cell-reactive compstatin analogs, cell-targeted compstatin analogs, and / or non-targeted compstatin analogs (e.g., long-acting non-targeted compstatin analogs) are used to protect blood cells from complement-mediated damage. Blood cells may be any of the cellular components of blood, e.g., red blood cells (RBCs), white blood cells (WBCs), and / or platelets. In one embodiment, cell-targeted compstatin analogs are targeted against target molecules such as glycophorin or band 3 exposed on the cell surface of RBCs. Many disorders are due to complement-mediated damage to blood cells. Such disorders may result from deficiencies or abnormalities in one or more intracellular or soluble CRPs in a patient, e.g., (a) mutations in genes encoding such proteins; (b) mutations in genes necessary for the production or proper function of one or more CRPs; and / or (c) the presence of one or more autoantibodies against CRP. Complement-mediated RBC lysis can result from the presence of autoantibodies against RBC antigens, which can be caused by a variety of factors (often idiopathic). Patients with such mutations in the gene encoding CRP and / or antibodies against CRP or their own RBCs are at increased risk of disorder involving complement-mediated RBC damage. Patients who have experienced one or more episodes of symptoms characteristic of the disorder are at increased risk of recurrence.

[0288] Paroxysmal nocturnal hemoglobinuria (PNH) is a relatively rare disorder, including acquired hemolytic anemia, characterized by complement-mediated intravascular hemolysis, hemoglobinuria, bone marrow failure, and a tendency to form blood clots. It is estimated that 16 people per million worldwide are affected, and while it can occur at any age and gender, it is more common in young adults (Bessler, M. & Hiken, J., Hematology Am Soc Hematol Educ Program, 104-110 (2008); Hillmen, P. Hematology Am Soc Hematol Educ Program, 116-123 (2008)). PNH is a chronic wasting disease interrupted by acute hemolysis, resulting in significant pathological conditions and a short life expectancy. Many patients experience abdominal pain, dysphagia, erectile dysfunction, and pulmonary hypertension in addition to anemia, and are at increased risk of renal failure and thromboembolism.

[0289] PNH was first described as a distinct condition in the 1800s, but the cause of hemolysis in PNH was not firmly established until the 1950s, when an alternative pathway for complement activation was discovered (Parker CJ. Paroxysmal nocturnal hemoglobinuria: an historical overview. Hematology Am Soc Hematol Educ Program. 93-103 (2008)). CD55 and CD59 are normally bound to the cell membrane via glycosylphosphatidylinositol (GPI) anchors (glycolipid structures that anchor specific proteins to the cell membrane). PNH results from non-malignant clonal proliferation of hematopoietic stem cells that have undergone somatic mutations in the PIGA gene, which encodes a protein involved in GPI anchor synthesis (Takeda J, et al. Deficiency of the GPI anchor caused by a somatic mutation of the PIG-A gene in paroxysmal nocturnal hemoglobinuria. Cell. 73: 703-711 (1993)). The offspring of these stem cells lack GPI-binding proteins, including CD55 and CD59. This deficiency makes the cells more susceptible to complement-mediated RBC lysis. Flow cytometry analysis using antibodies against GPI-binding proteins is commonly used for diagnosis. This analysis can detect the deficiency of GPI-binding proteins on the cell surface and reveal the degree of deficiency and the percentage of affected cells (Brodsky RA. Advances in the diagnosis and therapy of paroxysmal nocturnal hemoglobinuria. Blood Rev. 22(2): 65-74 (2008)). In PNHIII-type RBCs, GPI-binding proteins are completely deficient, resulting in increased sensitivity to complement, while in PNHII-type RBCs, some GPI-binding proteins are deficient, resulting in relatively lower sensitivity to complement.FLAER is a fluorescently labeled inactive variant of proaerolysin (a bacterial toxin that binds to a GPI anchor) and is increasingly used in conjunction with flow cytometry for the diagnosis of PNH. If binding of FLAER to granulocytes is not confirmed, PNH is diagnosed. In one embodiment, a cell-reactive compstatin analog protects PNH RBCs from C3b deposition.

[0290] In one embodiment, a cytoreactive, long-acting, or targeted compstatin analog is administered to a subject suffering from atypical hemolytic syndrome (aHUS). aHUS is a chronic disorder characterized by microangiogenic hemolytic anemia, thrombocytopenia, and acute renal failure, often caused by abnormal complement activation due to mutations in genes encoding complement regulatory proteins (Warwicker, P., et al.. Kidney Int 53, 836-844 (1998); Kavanagh, D. & Goodship, T. Pediatr Nephrol 25, 2431-2442 (2010)). Mutations in the complement H factor (CFH) gene are the most common genetic abnormality in aHUS patients, and 60-70% of these patients die within one year of disease onset or progress to end-stage renal failure (Kavanagh & Goodship, supra.). Other mutations described include those in factor I, factor B, C3, factor H-related proteins 1-5, and thrombomodulin. Other causes of aHUS include autoantibodies against complement regulatory proteins such as CFH. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is administered to a subject confirmed to have mutations in factor I, factor B, C3, factor H-related proteins 1-5, or thrombomodulin, or to a subject confirmed to have antibodies against complement regulatory proteins, such as CFH or converterase (e.g., C3Nef), or complement factors.

[0291] Complement-mediated hemolysis is seen in a diverse group of other conditions, including autoimmune hemolytic anemia associated with antibodies that bind to red blood cells (RBCs) and induce complement-mediated hemolysis. For example, such hemolysis can occur in primary chronic cold agglutinin disease, warm-antibody autoimmune hemolytic anemia (wAIHA), and certain reactions to drugs and other foreign substances (Berentsen, S., et al., Hematology 12, 361-370 (2007); Rosse, WF, Hillmen, P. & Schreiber, AD Hematology Am Soc Hematol Educ Program, 48-62 (2004)). In one embodiment, a cell-reactive compstatin analog is administered to a subject suffering from or at risk of chronic cold agglutinin disease. In one embodiment of the present invention, a cell-reactive compstatin analog is administered to a subject suffering from or at risk of wAIHA. In another embodiment, cell-reactive compstatin analogs were used to treat subjects with or at risk of HELLP syndrome, a disorder defined by hemolysis, elevated liver enzymes, and decreased platelet count, and caused in at least some patients by mutations in complement regulatory proteins (Fakhouri, F., et al., 112: 4542-4545 (2008)).

[0292] In another embodiment, cell-reactive compstatin analogs are used to protect RBCs or other cellular components of blood transfused to a subject. Some examples of such uses are further described below. As described above, targeted compstatin analogs and / or long-acting compstatin analogs can be used in the above methods for inhibiting complement-mediated hemolysis and / or RBC injury. In one embodiment, PNH or aHUS is treated with a long-acting compstatin analog containing a (CH2CH2O) moiety.

[0293] B. Transplantation Transplantation is an increasingly important therapeutic method that provides a means of replacing organs and tissues damaged by trauma, disease, or other conditions. The kidneys, liver, lungs, pancreas, and heart are among the organs that have been successfully transplanted. Tissues that are frequently transplanted include bone, cartilage, tendons, cornea, skin, heart valves, and blood vessels. Islet or islet cell transplantation is a promising treatment method for diabetes, such as type 1 diabetes. For the purposes of this invention, an organ, tissue, or cell (or group of cells) that is transplanted, has been transplanted, or has already been transplanted may be referred to as a “graft.” For the purposes of this specification, a blood transfusion is considered a “graft.”

[0294] In transplantation, the graft is exposed to a variety of events and stimuli that can cause graft dysfunction and potentially contribute to transplant failure. For example, ischemia-reperfusion (I / R) injury is a common and significant cause of morbidity and death in many grafts (especially solid organs) and can be a major determinant of graft survival. Transplant rejection is one of the major risks of transplantation between genetically different individuals and can lead to transplant failure, necessitating the removal of the graft from the recipient.

[0295] In one embodiment of the present invention, cell-reactive compstatin analogs, cell-targeted compstatin analogs, and / or long-acting compstatin analogs are used to protect grafts from complement-mediated injury. Cell-reactive compstatin analogs react with and covalently bind to graft cells, inhibiting complement activation. Cell-targeted compstatin analogs bind to target molecules in the graft (e.g., those expressed by graft endothelial cells or other cells), inhibiting complement activation. Target molecules may be, for example, molecules whose expression is induced or stimulated by stimuli such as injury or inflammation, molecules that can be recognized as "non-self" by the recipient, or molecules containing carbohydrate heteroantigens commonly found in humans, such as blood group antigens or heterogeneous antigens against which antibodies exist, such as alpha-gal epitopes. In one embodiment, reduced complement activation may be demonstrated by a decrease in the mean C4d deposition in blood vessels of grafts in contact with a compstatin analog, such as a cell-reactive compstatin analog, compared to the mean C4d deposition level in grafts not in contact with the compstatin analog (e.g., for a subject consistent with respect to the graft and other treatments received).

[0296] In various embodiments of the present invention, the graft can be contacted with a cell-reactive, long-acting, or targeted compstatin analog before, during, and / or after transplantation. For example, before transplantation, the graft removed from the donor can be contacted with a liquid containing a cell-reactive, long-acting, or targeted compstatin analog. For example, the graft can be immersed in the solution and / or perfused with the solution. In another embodiment, the cell-reactive, long-acting, or targeted compstatin analog is administered to the donor before the graft is removed. In one embodiment, the cell-reactive, long-acting, or targeted compstatin analog is administered to the recipient during and / or after the graft is introduced. In one embodiment, the cell-reactive, long-acting, or targeted compstatin analog is locally delivered to the transplanted graft. In one embodiment, the cell-reactive compstatin analog is administered systemically, for example, by intravenous administration.

[0297] The present invention provides a composition comprising an isolated graft (a) and a cell-reactive, long-acting, or targeted compstatin analog (b). In one embodiment, the composition further comprises a solution suitable for contacting a graft (e.g., an organ), such as an isolated graft removed from a donor and awaiting transplantation to a recipient (e.g., suitable for rinsing, washing, immersion, perfusion, maintenance, or storage). In one embodiment, the present invention provides a composition comprising a solution (a) suitable for contacting a graft (e.g., an organ) and a cell-reactive, long-acting, or targeted compstatin analog (b). The solution may be any solution that is physiologically acceptable to the graft (e.g., has an appropriate osmotic composition and is non-cytotoxic), medically acceptable in terms of subsequently introducing the graft into the recipient (e.g., preferably sterile, or at least reasonably free of microorganisms or other contaminants), and compatible with cell-reactive compstatin analogs (i.e., does not impair the reactivity of the compstatin analog) or compatible with long-acting or targeted compstatin analogs. In one embodiment, the solution is any solution known in the art for any such purpose. In one embodiment, the solution is Marshall's solution or a high-osmotic citrate solution (Soltran®, Baxter Healthcare), the University of Wisconsin (UW) solution (ViaSpan®, Bristol Myers Squibb), a histidine-tryptophan-ketoglutaric acid (HTK) solution (Custodial®, Kohler Medical Limited), EuroCollins (Fresenius) and Celsior® (Sangstat Medical), Polysol, IGL-1, or AQIX® RS-1. Naturally, other solutions within the range of physiologically acceptable compositions may be used, for example, solutions containing the same or nearly the same components at the same or different concentrations. In one embodiment, the solution does not contain components that are expected to elicit a significant reaction from the cell-reactive compstatin analog, and any solution can be modified or designed to not contain such components.In one embodiment, the cell-reactive compstatin analog may be present in a graft-compatible solution at a concentration of, for example, 0.01 to 100 mg / ml, or added to the solution until such a concentration is reached.

[0298] In one embodiment, the present invention provides a kit comprising a cell-reactive, long-acting, or targeted compstatin analog (a) and a graft-compatible solution or its solid (e.g., powder) component (b). The cell-reactive, long-acting, or targeted compstatin analog may be provided in solid form (e.g., powder) or at least partially dissolved in solution. In one embodiment, the cell-reactive, long-acting, or targeted compstatin analog and / or the graft-compatible solution are provided in predetermined quantities so as to obtain a solution of a concentration suitable for contacting the graft with the cell-reactive, long-acting, or targeted compstatin analog. In many embodiments, the cell-reactive, long-acting, or targeted compstatin analog and the graft-compatible solution or its solid (e.g., powder) component are housed in separate containers within the kit. In one embodiment, the cell-reactive compstatin analog and the components of the graft-compatible solution are housed together in separate containers or mixed and provided in solid (e.g., powder) form. In one embodiment, the kit includes instructions for use, e.g., instructions for adding a cell-reactive, long-acting, or targeted compstatin analog to a graft-compatible solution and / or instructions for contacting the graft with the cell-reactive compstatin analog. Optionally, the kit includes a label approved by a government agency relating to the regulation of products used in transplantation, cell therapy, and / or blood transfusion.

[0299] The present invention further provides a method for covalently bonding a compstatin analog to an isolated graft, comprising contacting the isolated graft with a cell-reactive compstatin analog. The present invention further provides an isolated graft to which a compstatin analog is covalently bonded. The isolated graft typically has a large number of compstatin analog molecules bound to it. In one embodiment, the graft is or contains a solid organ such as a kidney, liver, lung, pancreas, or heart. In one embodiment, the graft is or contains bone, cartilage, fascia, tendon, ligament, cornea, sclera, pericardium, skin, heart valve, blood vessel, amnion, or dura mater. In one embodiment, the graft comprises multiple organs such as a cardiopulmonary or pancreatic-kidney graft. In one embodiment, the graft comprises less than an entire organ or tissue. For example, the graft may comprise a portion of an organ or tissue, such as a liver lobe, a portion of blood vessels, a skin flap, or a heart valve. In one embodiment, the graft comprises a preparation comprising isolated cells or isolated tissue fragments isolated from the original tissue but retaining at least a portion of the tissue structure, such as pancreatic islets. In one embodiment, the preparation comprises a blood product containing isolated cells that are not bound to each other via connective tissue, such as hematopoietic stem cells or progenitor cells derived from peripheral blood and / or umbilical cord blood or whole blood, or any cells such as red blood cells (RBCs) or platelets. In one embodiment, the graft is obtained from a deceased donor (e.g., a “brain-dead donor” (DBD donor) or a “cardiac-dead donor” donor). In one embodiment, depending on the specific type of graft, the graft is obtained from a living donor. Types of grafts that can often be obtained from living donors without imposing undue risk to the donor and without deviating from legitimate medical practice include, for example, kidneys, parts of the liver, and blood cells.

[0300] In some embodiments, the transplant is xenotransplantation (i.e., the donor and recipient belong to different species). In some embodiments, the transplant is autotransplantation (i.e., transplantation from one part of the body to another within the same individual). In some embodiments, the transplant is syngeneic transplantation (i.e., the donor and recipient are genetically matched). In most embodiments, the transplant is allogeneic transplantation (i.e., the donor and recipient belong to the same species but are genetically different). In allogeneic transplantation, the donor and recipient may or may not be genetically related (e.g., members of the same family). Typically, the donor and recipient have compatible blood types (at least ABO compatibility, and possibly compatibility in Rh, Kell, and / or other blood cell antigens). Because the presence of alloantibodies can cause hyperacute rejection (i.e., rejection that begins almost immediately, e.g., within minutes, upon contact of the graft with the recipient's blood), the recipient's blood may be screened for alloantibodies against the graft and / or against the recipient and donor. The complement-dependent cell-mediated cytotoxicity (CDC) assay can be used to screen for anti-HLA antibodies in a target serum. The serum is incubated with lymphocytes of a known HLA phenotype. If the serum contains antibodies against the HLA molecules on the target cells, complement-mediated lysis and subsequent cell death will occur. Using a selected group of target cells, the specificity of the detected antibodies can be determined. Other useful techniques for determining the presence of anti-HLA antibodies and, if applicable, their HLA specificity include ELISA assays, flow cytometry assays, and microbead array technology (e.g., Luminex technology). Methodologies for performing these assays are well-known, and a variety of kits for performing them are commercially available.

[0301] In some embodiments, cell-reactive, long-acting, or targeted compstatin analogs suppress complement-mediated rejection. For example, in some embodiments, cell-reactive, long-acting, or targeted compstatin analogs suppress hyperacute rejection. Hyperacute rejection is caused, at least in part, by antibody-mediated activation of the recipient complement system via the classical pathway and the resulting MAC deposition in the graft. Hyperacute rejection is usually due to the presence of pre-existing antibodies in the recipient that react with the graft. While it is desirable to strive to avoid hyperacute rejection through adequate pre-transplant matching, this is not always possible due to, for example, time and / or supply constraints. Furthermore, some recipients (e.g., patients who have received several blood transfusions, patients who have previously undergone transplants, women who have been pregnant several times) have many pre-formed antibodies that may include antibodies against antigens that are not normally tested, making it difficult or nearly impossible to confidently obtain a timely and suitable graft. Such patients are at higher risk of hyperacute rejection.

[0302] In one embodiment, cell-reactive, long-acting, or targeted compstatin analogs suppress acute rejection or transplant failure. “Acute rejection” as used here refers to rejection occurring at least 24 hours after transplantation, typically for at least several days to a week, and up to six months after transplantation. Acute antibody-mediated rejection (AMR) often involves a rapid increase in donor-specific alloantibodies (DSAs) in the first few weeks after transplantation. While we do not wish to be bound by any theory, it is possible that existing plasma cells and / or newly converted plasma cells from memory B cells play some role in the increased production of DSAs. Such antibodies can cause complement-mediated damage to the graft, which can be inhibited by contacting the graft with cell-reactive compstatin analogs. Again, while we do not wish to be bound by any theory, inhibiting complement activation in the graft can reduce leukocyte (e.g., neutrophil) infiltration, another cause of acute transplant failure.

[0303] In one embodiment, cell-reactive, long-acting, or targeted compstatin analogs inhibit complement-mediated I / R injury to grafts. As will be further discussed below, I / R injury can occur during reperfusion of tissues where blood supply is temporarily disrupted, as seen in transplanted organs. Mitigating I / R injury may reduce the likelihood or severity of acute graft failure and thus the likelihood of acute transplant failure.

[0304] In one embodiment, cell-reactive, long-acting, or targeted compstatin analogs suppress chronic rejection and / or chronic transplant failure. As used herein, “chronic rejection or chronic transplant failure” refers to rejection or failure that occurs at least six months after transplantation, for example, six months to one, two, three, four, five years or more after transplantation, often after the graft has functioned normally for several months to several years. This is due to a chronic inflammatory immune response to the graft. For the purposes of this specification, chronic rejection may include chronic allograft vasculopathy, a term used to refer to fibrosis of the internal blood vessels of the transplanted tissue. Because immunosuppressive therapy has reduced the incidence of acute rejection, chronic rejection has become more prominent as a cause of graft dysfunction and transplant failure. There is evidence that B-cell alloantibody production is a key factor in the development of chronic rejection and chronic transplant failure (Kwun J. and Knechtle SJ, Transplantation, 88(8): 955-61 (2009)). Early damage to grafts can be a contributing factor to chronic processes such as fibrosis, which can ultimately lead to chronic rejection. Therefore, suppressing such early damage using cell-reactive compstatin analogs may delay the onset of chronic graft rejection and / or reduce the likelihood or severity of chronic graft rejection.

[0305] In one embodiment, a long-acting compstatin analog is administered to the graft recipient to suppress graft rejection and / or graft failure.

[0306] C. Ischemia / Reperfusion Injury Ischemia-reperfusion (I / R) injury is a significant cause of tissue damage in post-traumatic conditions and other conditions involving transient interruption of blood flow, such as myocardial infarction, stroke, severe infection, vascular disease, aneurysm repair, cardiopulmonary bypass, and transplantation.

[0307] In traumatic situations, systemic hypoxemia, hypotension, and localized disruption of blood supply due to contusions, compartment syndrome, and vascular injury lead to ischemia, damaging metabolically active tissues. Restoration of blood supply often triggers a strong systemic inflammatory response, which is more harmful than the ischemia itself. When reperfusion occurs at the ischemic site, locally produced and released factors enter the circulatory system and reach distant sites, sometimes causing significant injury to organs that were not originally affected by ischemic injury, such as the lungs and intestines, leading to single- and multi-organ dysfunction. Complement activation occurs immediately after reperfusion and is a crucial mediator of post-ischemic injury, directly and through its chemotactic and stimulant effects on neutrophils. All three major complement pathways are activated and act cooperatively or independently, contributing to I / R-related adverse events affecting multiple organ systems. In one embodiment of the present invention, a cellularly reactive, long-acting, or targeted compstatin analog is administered to a subject who has recently suffered trauma, such as systemic hypoxemia, hypotension, and / or local disruption of blood supply, which puts the subject at risk of I / R injury. In one embodiment, the cellularly reactive compstatin analog is administered intravascularly, optionally into the blood vessel supplying the injured body part, or directly to the body part. In one embodiment, the subject is a patient with spinal cord injury, traumatic brain injury, burns, and / or hemorrhagic shock.

[0308] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is administered to the target before, during, or after a surgical procedure, such as a surgical procedure that is expected to temporarily block blood flow to a tissue, organ, or part of the body. Examples of such procedures include cardiopulmonary bypass, angioplasty, heart valve repair / replacement, aneurysm repair, or other vascular surgery. The cell-reactive compstatin analog may be administered before, after, and / or during the time overlapping with the surgical procedure.

[0309] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is administered to a subject suffering from MI, thromboembolic stroke, deep vein thrombosis, or pulmonary embolism. The cell-reactive compstatin analog may be administered in combination with a thrombolytic agent such as tissue plasminogen activator (tPA) (e.g., alteplase, reteplase, tenecteplase), anistreplase (Eminase), streptokinase (Kabikinase, Streptase), or urokinase (Abbokinase). The cell-reactive, long-acting, or targeted compstatin analog may be administered before, after, and / or during the time overlap with the thrombolytic agent.

[0310] In one embodiment, cell-reactive, long-acting, or targeted compstatin analogs are administered to treat I / R injury.

[0311] D. Other complement-mediated disorders In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is introduced intraocularly to treat an ocular disorder such as age-related macular degeneration (AMD), diabetic retinopathy, glaucoma, or uveitis. For example, a cell-reactive, long-acting, or targeted compstatin analog is introduced intravitreally (e.g., by intravitreal injection) to treat a subject with AMD or at risk of developing AMD. In one embodiment, AMD is neovascular (exudative) AMD. In another embodiment, AMD is dry AMD. As will be recognized by those skilled in the art, dry AMD includes geographic atrophy (GA), intermediate AMD, and early AMD. In one embodiment, a subject with GA is treated to delay or halt the progression of the disease. For example, in one embodiment, treatment of a subject with GA reduces the rate of retinal cell death. The reduction in the rate of retinal cell death can be demonstrated by a reduction in the growth rate of GA lesions in patients treated with LACA compared to a control (e.g., a patient injected with sham). In one embodiment, the subject has intermediate-stage AMD. In another embodiment, the subject has early-stage AMD. In another embodiment, the subject having intermediate-stage or early-stage AMD is treated to delay or halt the progression of the disease. For example, in one embodiment, treatment of the subject having intermediate-stage AMD may delay or prevent progression to a progressive form of AMD (neovascular AMD or GA). In another embodiment, treatment of the subject having early-stage AMD may delay or prevent progression to intermediate-stage AMD. In another embodiment, the eye has both GA and neovascular AMD. In another embodiment, the eye has GA but not exudative AMD. In another embodiment, a cell-reactive, long-acting, or targeted compstatin analog is injected intravitreally for the treatment of glaucoma, uveitis (e.g., posterior uveitis), or diabetic retinopathy. In another embodiment, a cell-reactive, long-acting, or targeted compstatin analog is introduced into the anterior chamber to treat, for example, anterior uveitis.

[0312] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects suffering from or at risk of autoimmune diseases, such as autoimmune diseases mediated by antibodies against at least one or more autoantigens.

[0313] Cell-reactive, long-acting, or targeted compstatin analogs can be introduced, for example, into the synovial fluid of a subject suffering from arthritis (e.g., rheumatoid arthritis). Naturally, these may also be administered systemically.

[0314] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects who have suffered from or are at risk of intracerebral hemorrhage.

[0315] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects who have myasthenia gravis or are at risk of developing myasthenia gravis.

[0316] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects with or at risk of developing neuromyelitis optica (NMO).

[0317] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects suffering from or at risk of membranoproliferative glomerulonephritis (MPGN), such as type I MPGN, type II MPGN, or type III MPGH.

[0318] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects suffering from or at risk of developing neurodegenerative diseases. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects suffering from or at risk of developing neuropathic pain. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects suffering from or at risk of developing rhinosinusitis or nasal polyps. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects suffering from or at risk of developing cancer. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects suffering from or at risk of developing sepsis. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat subjects who have or are at risk of developing adult respiratory distress syndrome.

[0319] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat a subject who has or is at risk of anaphylaxis or an infusion reaction. For example, in one embodiment, the subject may be pre-treated before, during, or after administration of a drug or solvent that may cause anaphylaxis or an infusion reaction. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat a subject who is at risk of or is experiencing anaphylaxis due to food (e.g., peanuts, shellfish, or other food allergens) or insect stings (e.g., bees, wasps).

[0320] In various embodiments of the present invention, cell-reactive, long-acting, or targeted compstatin analogs may be administered topically or systemically.

[0321] In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat respiratory diseases, such as asthma, chronic obstructive pulmonary disease (COPD), or idiopathic pulmonary fibrosis. In various embodiments, for example, a cell-reactive, long-acting, or targeted compstatin analog may be administered into the airways, for example, as a dry powder or by inhalation via spray, or by injection, for example, intravenously, intramuscularly, or subcutaneously. In one embodiment, a cell-reactive, long-acting, or targeted compstatin analog is used to treat severe asthma, such as asthma that is not adequately controlled by bronchodilators and / or inhaled corticosteroids.

[0322] In one embodiment, a method for treating a complement-mediated disorder, such as a chronic complement-mediated disorder, is provided, comprising administering a long-acting complement inhibitor to a subject requiring treatment of the disorder. The long-acting compstatin analog may be any of the long-acting compstatin analogs described herein in various embodiments. In one embodiment, a method for treating a Th17-related disorder is provided, comprising administering a long-acting complement inhibitor to a subject requiring treatment of the disorder.

[0323] In some embodiments, a “chronic disorder” is a disorder that lasts for at least three months and / or is recognized as a chronic disorder in this art. In many embodiments, a chronic disorder lasts for at least six months, for example, at least one year or longer, for example, permanently. Those skilled in the art will recognize that at least some findings of various chronic disorders may be intermittent and / or their severity may worsen and remission over time. Chronic disorders are progressive and may, for example, become more severe or more widespread over time. Many chronic complement-mediated disorders are described here. A chronic complement-mediated disorder may be any chronic disorder in which complement activation (e.g., excessive or inappropriate complement activation) is involved as, for example, the cause and / or at least some causative factor. For convenience, disorders may be classified by referring to the organ or system in which they occur particularly frequently in the affected subject. It is recognized that many disorders may occur in multiple organs or systems, and such classification is not limiting in any way. Furthermore, many findings (e.g., symptoms) may occur in subjects with any of many different disorders. Non-limiting information regarding the disorders discussed here can be found, for example, in standard textbooks of internal medicine, such as the Cecil Textbook of Medicine (e.g., 23rd edition), Harrison's Principles of Internal Medicine (e.g., 17th edition), and / or standard textbooks focusing on specific medical areas, specific body systems or organs, and / or specific disorders.

[0324] In one embodiment, chronic complement-mediated disorder is Th2-related disorder. Th2-related disorder as used herein is a disorder characterized by an excessive number and / or excessive or inappropriate activity of Th2 subtype CD4+ helper T cells ("Th2 cells") in the body or a part of the body, e.g., at least one tissue, organ, or structure. For example, in a disordered tissue, organ, or structure, Th2 cells may be dominant over Th1 subtype CD4+ helper T cells ("Th1 cells"). As is known in this art, Th2 cells typically secrete characteristic cytokines such as interleukin-4 (IL-4), interleukin-5 (IL-5), and interleukin-13 (IL-13), while Th1 cells typically secrete interferon-γ (IFN-γ) and tumor necrosis factor β (TNFβ). In one embodiment, Th2-related dysfunction is characterized, for example, by the production and / or quantity of excessive IL-4, IL-5, and / or IL-13 in relation to IFN-γ and / or TNFβ in at least one tissue, organ, or structure.

[0325] In one embodiment, chronic complement-mediated disorders are Th17-related disorders. In one embodiment, as further described in PCT / US2012 / 043845, filed June 22, 2012, entitled “Method for the Treatment of Chronic Disorders with Complement Inhibitors,” complement activation and Th17 cells participate in a cycle involving dendritic cells and antibodies, contributing to the maintenance of a pathogenic immune microenvironment underlying a certain range of disorders. While we do not wish to be bound by any theory, the pathogenic immune microenvironment, once established, is autonomous and contributes to cellular and tissue damage. In one embodiment, long-acting compstatin analogs are useful for the treatment of Th17-related disorders.

[0326] The Th17-related disorder as used herein is a disorder characterized by an excessive number and / or excessive or inappropriate activity of CD4+ helper T cells of the Th17 subtype ("Th17 cells") in the body or a part of the body, e.g., at least one tissue, organ, or structure. For example, Th17 cells may be dominant over Th1 and / or Th2 cells in the affected tissue, organ, or structure. In some embodiments, the Th17 cell dominance is relative, e.g., the Th17 cell to Th1 cell ratio and / or Th17 cell to Th2 cell ratio are elevated relative to normal values. In some embodiments, the Th17 cell to regulatory T cell ratio (CD4) is elevated. + CD25 +Regulatory T cells, also known as “Treg cells,” are elevated compared to normal levels. Th17 cell formation and / or activation are promoted by various cytokines, e.g., interleukin-6 (IL-6), interleukin-21 (IL-21), interleukin-23 (IL-23), and / or interleukin-1β (IL-1β). Th17 cell formation includes the differentiation of precursor T cells, e.g., naive CD4+ T cells, into the Th17 phenotype and their subsequent differentiation into functional Th17 cells. In some embodiments, Th17 cell formation encompasses certain aspects of Th17 cell development, proliferation (increase), survival, and / or maturation. In some embodiments, Th17-related disorders are characterized by the excessive production and / or quantity of IL-6, IL-21, IL-23, and / or IL-1β. Th17 cells typically secrete characteristic cytokines such as interleukin-17A (IL-17A), interleukin-17F (IL-17F), interleukin-21 (IL-21), and interleukin-22 (IL-22). In one embodiment, Th17-related dysfunction is characterized by the overproduction and / or quantity of Th17 effector cytokines, e.g., IL-17A, IL-17F, IL-21, and / or IL-22. In one embodiment, the overproduction or quantity of cytokines is detectable in the blood. In one embodiment, the overproduction or quantity of cytokines is detectable locally, for example, in at least one tissue, organ, or structure. In one embodiment, Th17-related dysfunction is associated with a decrease in the number of Tregs and / or a decrease in the amount of Treg-related cytokines. In one embodiment, Th17 disorder is some kind of chronic inflammatory disease, and the term encompasses a range of discomfort characterized by autoperpetual immune invasion of various tissues, which appears to be dissociated from the initial invasion (which may be unknown) that causes the discomfort. In one embodiment, Th17-related disorder is some kind of autoimmune disease. Most, if not most, “chronic inflammatory diseases” are actually autoimmune diseases.Examples of Th17-related disorders include inflammatory skin diseases such as psoriasis and atopic dermatitis; systemic scleroderma and sclerosis; inflammatory bowel disease (IBD) (e.g., Crohn's disease and ulcerative colitis); Behçet's disease; dermatomyositis; polymyositis; multiple sclerosis (MS); dermatitis; meningitis; encephalitis; uveitis; osteoarthritis; lupus nephritis; rheumatoid arthritis (RA), Sjögren's syndrome, multiple sclerosis, vasculitis; central nervous system (CNS) inflammatory disorders, chronic hepatitis; chronic pancreatitis, glomerulonephritis; sarcoidosis; thyroiditis; pathological immune responses to tissue / organ transplantation (e.g., graft rejection); COPD, asthma, bronchiolitis, hypersensitivity pneumonitis, idiopathic pulmonary fibrosis (IPF), periodontitis, and gingivitis. In some embodiments, Th17 disorders are older known autoimmune diseases such as type 1 diabetes or psoriasis. In one embodiment, Th17-related disorder is age-related macular degeneration.

[0327] In one embodiment, chronic complement-mediated disorder is an IgE-related disorder. As used herein, “IgE-related disorder” is a disorder characterized by excessive and / or inadequate production and / or quantity of IgE, excessive or inadequate activity of IgE-producing cells (e.g., IgE-producing B cells or plasma cells) and / or excessive and / or inadequate activity of IgE-responsive cells such as eosinophils or mast cells. In one embodiment, the IgE-related disorder is characterized by high levels of total IgE and / or, in one embodiment, allergen-specific IgE in the plasma of the subject and / or locally.

[0328] In one embodiment, chronic complement-mediated disorder is characterized by the presence of autoantibodies and / or immune complexes in the body that can activate complement, for example, via the classical pathway. Autoantibodies can bind to, for example, autoantigens on cells or tissues in the body. In one embodiment, autoantibodies can bind to antigens such as those in blood vessels, skin, nerves, muscles, connective tissue, heart, kidneys, and thyroid gland. In one embodiment, the subject has neuromyelitis optica and produces autoantibodies against aquaporin 4 (e.g., IgG autoantibodies). In one embodiment, the subject has bullous pemphigoid and produces autoantibodies (e.g., IgG or IgE autoantibodies) against structural elements of hemiadhesive plaques (e.g., transmembrane collagen XVII (BP180 or BPAG2) and / or plakin family protein BP230 (BPAG1)). In another embodiment, the subject has polyneuropathy, such as chronic or acute polyneuropathy like multifocal motor neuropathy, and produces anti-ganglioside antibodies that bind to gangliosides present in nerve cell membranes. In yet another embodiment, chronic complement-mediated dysfunction is not characterized by autoantibodies and / or immune complexes.

[0329] In one embodiment, the chronic complement-mediated disorder is a respiratory disorder. In one embodiment, the chronic respiratory disorder is asthma or chronic obstructive pulmonary disease (COPD). In one embodiment, the chronic respiratory disorder is pulmonary fibrosis (e.g., idiopathic pulmonary fibrosis), radiation-induced lung injury, allergic bronchopulmonary aspergillosis, hypersensitivity pneumonitis (also known as allergic alveolitis), eosinophilic pneumonia, interstitial pneumonia, sarcoid, Wegener's granuloma, or obstructive bronchiolitis. In one embodiment, the present invention provides a method for treating a chronic respiratory disorder in a subject requiring treatment, e.g., asthma, COPD, pulmonary fibrosis, radiation-induced lung injury, allergic bronchopulmonary aspergillosis, hypersensitivity pneumonitis (also known as allergic alveolitis), eosinophilic pneumonia, interstitial pneumonia, sarcoid, Wegener's granuloma, or obstructive bronchiolitis, comprising administering a long-acting complement inhibitor to the subject requiring treatment of the disorder.

[0330] In one embodiment, the chronic complement-mediated disorder is allergic rhinitis, sinusitis, or nasal polyposis. In one embodiment, the present invention provides a method for treating allergic rhinitis, sinusitis, or nasal polyposis in a subject requiring treatment, comprising administering a long-acting complement inhibitor to the subject requiring treatment of the disorder.

[0331] In one embodiment, chronic complement-mediated disorders are disorders that manifest in the musculoskeletal system. Examples of such disorders include inflammatory joint conditions (e.g., arthritis such as rheumatoid arthritis or psoriatic arthritis, juvenile chronic arthritis, spondyloarthropathy, Reiter's syndrome, and gout). In one embodiment, musculoskeletal disorders produce symptoms such as pain, stiffness, and / or limited movement in the affected part of the body. Inflammatory myopathy includes dermatomyositis, polymyositis, and various others that are disorders of chronic muscle inflammation of unknown cause that cause muscle weakness. In one embodiment, chronic complement-mediated disorders are myasthenia gravis. In one embodiment, the present invention provides a method for treating any of the musculoskeletal disorders, comprising administering a long-acting complement inhibitor to a subject requiring treatment of the disorder.

[0332] In one embodiment, chronic complement-mediated disorders are disorders that manifest in the cutaneous system. Examples of such disorders include, for example, atopic dermatitis, psoriasis, bullous ...

Claims

1. A physiologically acceptable or pharmaceutical-grade composition comprising a cell-reactive compstatin analog.

2. The composition according to claim 1, which is physiologically acceptable.

3. The composition according to claim 1, which is a pharmaceutical-grade composition.

4. The composition according to claim 1, which is pharmaceutically acceptable for administration to humans.

5. The composition according to claim 1, wherein the cell-reactive compstatin analog comprises a cell-reactive functional group that can covalently bind to mammalian cells.

6. The composition according to claim 1, wherein the cell-reactive compstatin analog comprises a cell-reactive moiety containing a cell-reactive functional group.

7. The composition according to claim 1, wherein the compstatin analog comprises a cell-reactive functional group that reacts with a sulfidyl (SH) group to form a covalent bond.

8. The composition according to claim 1, wherein the compstatin analog comprises a cell-reactive functional group that reacts with an amine group to form a covalent bond.

9. The composition according to claim 1, wherein the cell-reactive compstatin analog comprises a maleimide group.

10. The composition according to claim 1, wherein the cell-reactive compstatin analog comprises a cell-reactive moiety, and the cell-reactive moiety comprises a cell-reactive functional group.

11. The composition according to claim 1, wherein the cell-reactive compstatin analog comprises a cell-reactive functional group, a compstatin analog moiety, and a binding moiety that separates the cell-reactive functional group from the compstatin analog moiety of the cell-reactive compstatin analog.

12. The composition according to claim 1, wherein the cell-reactive compstatin analog comprises an amino acid whose side chain includes a group of formula (NH)-R (where R represents a portion containing a cell-reactive functional group).

13. The composition according to claim 1, wherein the cell-reactive compstatin analog is a compound having the core sequence X'aa-Gln-Asp-Xaa-Gly (SEQ ID NO: 3), wherein X'aa and Xaa are compounds comprising cyclic peptides selected from Trp and Trp analogs, and the compound comprises a cell-reactive moiety.

14. The composition according to claim 1, wherein the compstatin analog is a compound having the core sequence X'aa-Gln-Asp-Xaa-Gly-X"aa (SEQ ID NO: 4), wherein X'aa and Xaa are each independently selected from Trp and Trp analogs, and X"aa comprises a cyclic peptide selected from His, Ala, monomethyl unbranched amino acids, Phe, Trp and Trp analogs, and the compound comprises a cell-reactive moiety.

15. The composition according to claim 1, wherein the peptide has the sequence X'aa1-X'aa2-X'aa3-X'aa-Gln-Asp-Xaa-Gly-X"aa-X"aa2-X"aa3-X"aa4-X"aa5 (SEQ ID NO: 5), and X'aa1, X'aa2, X'aa3, X"aa2, X"aa3-X"aa4 and X"aa5 correspond to amino acids at corresponding positions in compstatin, and the compound comprises a cell-reactive moiety.

16. The composition according to claim 1, wherein the compstatin analog has the sequence X'aa1-X'aa2-X'aa3-X'aa4-Gln-Asp-Xaa-Gly-X"aa1-X"aa2-X"aa3-X"aa4-X"aa5 (SEQ ID NO: 5), and the compound comprises a cyclic peptide selected from Trp and Trp analogs, where X'aa4 and Xaa are independently selected from amino acids and amino acid analogs, the peptide is cyclized via a bond between X'aa2 and X"aa4, and the compound comprises a cell-reactive moiety.

17. The composition according to claim 16, wherein X'aa2 and X''aa4 are Cys, and X''aa1 is optionally Ala or a monomethyl unbranched amino acid, and the compound comprises a cell-reactive moiety.

18. The composition according to claim 16, wherein X'aa2 and X"aa4 are Cys, one or more of X'aa1, X'aa2, X'aa3, X"aa2, X"aa3, X"aa4 and X"aa5 correspond to amino acids at the corresponding positions of compstatin, X"aa1 is Ala or a monomethyl unbranched amino acid, and the compound comprises a cell-reactive moiety.

19. The compstatin analog has the sequence Xaa1-Cys-Val-Xaa2-Gln-Asp-Xaa2 ＊ It has -Gly-Xaa3-His-Arg-Cys-Xaa4 (Sequence ID 6), in the formula, Xaa1 is Ile, Val, Leu, B １ - Ile, B １ -Val, B １ -Leu or Gly-Ile or B １ It is a dipeptide containing -Gly-Ile, B １ This represents the first blocking section, Xaa2 and Xaa2 ＊ These are independently selected from Trp and its analogues, Xaa3 is His, Ala or an analogue of Ala, Phe, Trp or an analogue of Trp, Xaa4 is a dipeptide selected from L-Thr, D-Thr, Ile, Val, Gly, Thr-Ala, and Thr-Asn, or a tripeptide containing Thr-Ala-Asn, and any of the carboxyl-terminal -OH of L-Thr, D-Thr, Ile, Val, Gly, Al, or Asn optionally forms a second blockage B ２ It has been replaced by, This compound contains a cyclic peptide in which two Cys residues are linked by a disulfide bond. The compound contains a cell-reactive moiety. The composition according to claim 1.

20. Xaa1 is a dipeptide containing Ile, Val, Leu, Ac-Ile, Ac-Val, Ac-Leu, Gly-Ile, or Ac-Gly-Ile. Xaa2 and Xaa2 ＊ each independently is selected from Trp and analogs of Trp, Xaa3 is His, Ala or an analogue of Ala, Phe, Trp or an analogue of Trp, Xaa4 is a dipeptide selected from L-Thr, D-Thr, Ile, Val, Gly, Thr-Ala, and Thr-Asn, or a tripeptide containing Thr-Ala-Asn, and any of the carboxyl-terminal -OH of L-Thr, D-Thr, Ile, Val, Gly, Al, or Asn may be -NH ２ It has been replaced by The composition according to claim 19.

21. The composition according to claim 19, wherein Xaa2 is an analog of Trp having increased hydrophobicity compared to Trp.

22. The composition according to claim 19, wherein Xaa2 is a substituted or unsubstituted bicyclic aromatic ring component or an analog of Trp comprising two or more substituted or unsubstituted monocyclic aromatic ring components.

23. Xaa2 ＊ The composition according to claim 19, wherein is an analog of Trp having an electronegative substituent on the indole ring and not having increased hydrophobicity compared to Trp.

24. Xaa2 ＊ The composition according to claim 19, wherein the compound is an analog of Trp comprising a lower alkoxy substituent or a lower alkyl substituent at the 1st or 5th position of tryptophan, or a halogen substituent at the 5th or 6th position of tryptophan.

25. Xaa2 ＊ Xaa2 is an analog of Trp containing a lower alkoxy substituent or a lower alkyl substituent at the 1st or 5th position of tryptophan, or a halogen substituent at the 5th or 6th position of tryptophan. ＊ The composition according to claim 19, wherein is Trp.

26. The composition according to claim 19, wherein the compstatin analog comprises a cyclic peptide having a sequence selected from the group consisting of SEQ ID NOs: 9 to 36 and a cell-reactive moiety.

27. The composition according to claim 19, wherein the compstatin analog comprises a cyclic peptide having a sequence selected from the group consisting of SEQ ID NOs: 14, 21, 28, 29, 32, 33, 34, or 36, and a cell-reactive moiety.

28. The composition according to claim 19, wherein the compstatin analog comprises a cyclic peptide having the sequence of SEQ ID NO: 28, 32, or 34.

29. The composition according to claim 1, wherein the compstatin analog has the sequence X'aa1-X'aa2-X'aa3-X'aa4-Gln-Asp-Xaa-Gly-X"aa1-X"aa2-X"aa3-X"aa4-X"aa5 (SEQ ID NO: 5), wherein X'aa4 and Xaa are compounds comprising cyclic peptides selected from Trp and Trp analogs, X'aa1, X'aa2, X'aa3, X"aa1, X"aa2, X"aa3, X"aa4 and X"aa5 are independently selected from amino acids and amino acid analogs, X'aa2 and X"aa4 are not Cys, the peptide is cyclized via a bond between X'aa2 and X"aa4, and the compound comprises a cell-reactive moiety.

30. The composition according to claim 29, wherein one or more of X'aa1, X'aa3, X"aa2, X"aa3, and X"aa5 correspond to amino acids at the corresponding positions of the peptide according to any one of claims 18 to 28, and X"aa1 is Ala or a monomethyl unbranched amino acid.

31. The composition according to claim 29, wherein one of X'aa2 and X''aa4 is an amino acid or amino acid analog having a side chain containing a primary or secondary amine, and the other of X'aa2 and X''aa4 is an amino acid or amino acid analog having a side chain containing a carboxylic acid group, and the bond is an amide bond.

32. The composition according to claim 29, wherein X'aa1, X'aa3, X"aa1, X"aa2, X"aa3, and X"aa5 correspond to amino acids at the corresponding positions of the cyclic peptide described in any of claims 18 to 28, and optionally one of X'aa2 and X"aa4 is an amino acid or amino acid analog having a side chain containing a primary or secondary amine, and the other of X'aa2 and X"aa4 is an amino acid or amino acid analog having a side chain containing a carboxylic acid group, and the bond is an amide bond.

33. The composition according to any one of claims 10 to 32, wherein the cyclic peptide is acetylated at the N-terminus, amidated at the C-terminus, or acetylated at the N-terminus and amidated at the C-terminus.

34. Isolated cells or organs containing covalently bound compstatin analogs.

35. The isolated cell or isolated organ according to claim 34, which is a human cell or a human organ.

36. The isolated cell or isolated organ according to claim 34, wherein the isolated cell is a blood cell or the isolated organ is a heart, kidney, liver, lung, or pancreas.

37. A method for reducing the susceptibility of a cell or organ to complement-dependent injury, comprising contacting the cell with a cell-reactive compstatin analog, wherein the cell-reactive compstatin analog covalently binds to the cell or organ.

38. The method according to claim 37, wherein the cell or organ is a human cell or a human organ.

39. The method according to claim 37, wherein the cells are blood cells or the organ is the heart, kidney, liver, lung, or pancreas.

40. The method according to claim 37, wherein the cells or organ are isolated cells or an isolated organ to be transplanted to a subject, and the method comprises contacting the cells or organ with the cell-reactive compstatin analog before transplantation.

41. The method according to claim 37, wherein the cells or organ are transplanted to a target, and the organ is brought into contact with the cell-reactive compstatin analog after transplantation.

42. The method according to claim 37, comprising perfusing the organ with a liquid containing the cell-reactive compstatin analog.

43. The method according to claim 37, comprising bringing the cells or the organ into contact with the cell-reactive compstatin analog when the organ is transplanted into a target.

44. The method according to claim 37, comprising transplanting the cells or organs into the subject and then administering the cell-reactive compstatin analog to the subject.

45. The method according to claim 37, comprising administering the cell-reactive compstatin analog to the target after transplanting the organ to the target, wherein the cell-reactive compstatin analog is administered locally to the transplanted organ.

46. The method according to claim 37, wherein the cells or organs are transplanted into or to be transplanted into a subject at high risk of developing a hyperacute or acute complement-mediated transfusion reaction or complement-mediated organ rejection reaction.

47. A method for treating a subject requiring treatment for complement-mediated dysfunction, comprising administering a cell-reactive compstatin analog to the subject.

48. The method according to claim 47, wherein the cell-reactive compstatin analog is administered locally to a site at risk of complement-mediated injury or a site where complement-mediated injury has occurred.

49. The method according to claim 47, wherein the disorder causes complement-mediated injury in red blood cells, and the cell-reactive compstatin analog is administered intravascularly.

50. The method according to claim 47, wherein the subject has a defect in complement control.

51. The method according to claim 47, wherein the subject requires treatment for transplant rejection.

52. The method according to claim 47, wherein the subject requires treatment for ischemia / reperfusion injury.

53. The method according to claim 47, wherein the subject requires treatment for hemolytic anemia.

54. A long-acting compstatin analog comprising one or more compstatin analog moieties and a clearance reduction moiety (CRM) containing a polymer with a molecular weight of 10 kilodaltons (kD) to 45 kD.

55. The long-acting compstatin analog according to claim 54, comprising a polymer with a molecular weight of 20 kilodaltons (kD) to 45 kD as the CRM.

56. A long-acting compstatin analog according to claim 54 or 55, comprising a polymer with a molecular weight of 30 kilodaltons (kD) as a clearance reduction portion.

57. A long-acting compstatin analog according to claim 54 or 55, comprising a polymer with a molecular weight of 40 kilodaltons (kD) as a clearance reduction portion.

58. A long-acting compstatin analog according to any one of claims 54 to 57, wherein the polymer comprises polyethylene glycol (PEG).

59. A long-acting compstatin analog according to any one of claims 54 to 58, wherein the polymer contains linear PEG.

60. A long-acting compstatin analog according to any one of claims 54 to 58, wherein the polymer contains branched PEG.

61. The long-acting compstatin analog according to claim 59, wherein the polymer comprises a linear PEG and a compstatin analog moiety bound to each end of the linear PEG.

62. The long-acting compstatin analog according to claim 60, wherein the polymer comprises branched PEG having 3 to 10 branches.

63. The long-acting compstatin analog according to claim 62, wherein the polymer comprises a branched PEG having 3 to 10 branches, and at least about 50% of the branches have a compstatin analog moiety bound thereto.

64. The long-acting compstatin analog according to claim 63, wherein the polymer comprises a branched PEG having 3 to 10 branches, and at least about 75% of the branches have a compstatin analog moiety bound thereto.

65. A long-acting compstatin analog according to any one of claims 54 to 57, wherein the polymer contains human serum albumin.

66. A long-acting compstatin analog according to any one of claims 54 to 65, comprising 2 to 10 compstatin analog moieties.

67. A long-acting compstatin analog according to any one of claims 54 to 66, comprising 2 to 100 compstatin analog moieties.

68. A long-acting compstatin analog according to any one of claims 54 to 67, wherein the plasma half-life after intravenous injection into primates is at least two days.

69. A long-acting compstatin analog according to any one of claims 54 to 68, wherein the plasma half-life after intravenous injection into primates is at least 3 days.

70. A long-acting compstatin analog according to any one of claims 54 to 69, wherein the plasma half-life after intravenous injection into primates is at least 4 days.

71. A long-acting compstatin analog according to any one of claims 54 to 70, having the same amino acid sequence but lacking the clearance-reducing portion, and possessing at least about 20% of the molar activity of a corresponding compstatin analog.

72. A long-acting compstatin analog according to any one of claims 54 to 71, having molar activity of at least about 30% of the activity of the corresponding compstatin analog that does not include the clearance reduction portion.

73. A long-acting compstatin analog according to any one of claims 54 to 72, comprising multiple compstatin analog moieties and having a molar activity approximately equal to the sum of the activities of the compstatin analog moieties.

74. A long-acting compstatin analog according to any one of claims 54 to 73, wherein the terminal phase half-life is at least five times that of a corresponding compstatin analog that does not contain an equivalent amount of the CRM.

75. C ｍａｘ However, the long-acting compstatin analog according to any one of claims 54 to 74 is at least 10 times that of the corresponding compstatin analog which does not contain an equivalent amount of the clearance reduction portion.

76. A long-acting compstatin analog according to any one of claims 54 to 75, wherein the plasma half-life after subcutaneous injection into primates is at least two days.

77. The long-acting compstatin analog according to claim 76, wherein the plasma half-life after subcutaneous injection into primates is at least 3 days.

78. The long-acting compstatin analog according to claim 76, wherein the plasma half-life after subcutaneous injection into primates is at least 4 days.

79. A long-acting compstatin analog according to any one of claims 76 to 78, having molar activity of at least about 20% of the activity of the corresponding compstatin analog that does not contain a clearance reduction portion.

80. A long-acting compstatin analog according to any one of claims 76 to 79, having at least about 30% of the molar activity of a compstatin analog containing the same amino acid sequence but without the clearance-reducing portion.

81. A long-acting compstatin analog according to any one of claims 76 to 79, comprising a plurality of compstatin analog moieties and having a molar activity of at least about 10% of the total activity of the compstatin analog moieties.

82. C ｍａｘ The long-acting compstatin analog according to claim 76, wherein the amount is at least 10 times that of the corresponding compstatin analog that does not contain an equivalent amount of the clearance reduction portion.

83. At least about 30% of the activity and at least 10 times that of the corresponding compstatin analog that does not contain an equivalent amount of the clearance reduction portion. ｍａｘ A long-acting compstatin analog according to any one of claims 76 to 82, having a plasma half-life of at least 3 days.

84. The long-acting compstatin analog according to any one of claims 54 to 83, wherein the compstatin analog portion has the core sequence X'aa-Gln-Asp-Xaa-Gly (SEQ ID NO: 3), and in the formula, X'aa and Xaa comprise a cyclic peptide selected from Trp and an analog of Trp.

85. A long-acting compstatin analog according to any one of claims 54 to 83, wherein the compstatin analog portion has the core sequence X'aa-Gln-Asp-Xaa-Gly-X"aa (SEQ ID NO: 4), in which X'aa and Xaa are each independently selected from Trp and Trp analogs, and X"aa comprises a cyclic peptide selected from His, Ala, monomethyl unbranched amino acids, Phe, Trp and Trp analogs.

86. The long-acting compstatin analog according to any one of claims 54 to 83, wherein the compstatin analog portion comprises a cyclic peptide having the sequence X'aa1-X'aa2-X'aa3-X'aa4-X'aa5 (SEQ ID NO: 5), and X'aa1, X'aa2, X'aa3, X'aa2, X'aa3-X'aa4 and X'aa5 correspond to amino acids at the corresponding positions in compstatin.

87. A long-acting compstatin analog according to any one of claims 54 to 83, wherein the compstatin analog moiety has the sequence X'aa1-X'aa2-X'aa3-X'aa4-Gln-Asp-Xaa-Gly-X"aa1-X"aa2-X"aa3-X"aa4-X"aa5 (SEQ ID NO: 5), wherein X'aa4 and Xaa comprise a cyclic peptide selected from Trp and Trp analogs, and X'aa1, X'aa2, X'aa3, X"aa1, X"aa2, X"aa3, X"aa4 and X"aa5 are independently selected from amino acids and amino acid analogs, and the peptide is cyclized via a bond between X'aa2 and X"aa4.

88. A long-acting compstatin analog according to any one of claims 54 to 83, wherein X'aa2 and X''aa4 are Cys, and X''aa1 is optionally Ala or a monomethyl unbranched amino acid.

89. A long-acting compstatin analog according to any one of claims 54 to 83, wherein X'aa2 and X"aa4 are Cys, one or more of X'aa1, X'aa2, X'aa3, X"aa2, X"aa3, X"aa4 and X"aa5 correspond to the amino acids at the corresponding positions in compstatin, and X"aa1 is Ala or a monomethyl unbranched amino acid.

90. The compstatin analog portion is Sequence Xaa1-Cys-Val-Xaa2-Gln-Asp-Xaa2 ＊ -Gly-Xaa3-His-Arg-Cys-Xaa4(Sequence ID 6) It has, in the formula, Xaa1 is Ile, Val, Leu, B １ - Ile, B １ -Val, B １ -Leu or Gly-Ile or B １ It is a dipeptide containing -Gly-Ile, B １ This represents the first blocking section, Xaa2 and Xaa2 ＊ These are independently selected from Trp and its analogues, Xaa3 is His, Ala or an analogue of Ala, Phe, Trp or an analogue of Trp, Xaa4 is a dipeptide selected from L-Thr, D-Thr, Ile, Val, Gly, Thr-Ala, and Thr-Asn, or a tripeptide containing Thr-Ala-Asn, and any of the carboxyl-terminal -OH of L-Thr, D-Thr, Ile, Val, Gly, Al, or Asn optionally forms a second blockage B ２ It has been replaced by, A cyclic peptide in which two Cys residues are linked by a disulfide bond. A long-acting compstatin analog according to any one of claims 54 to 83, comprising the above.

91. Xaa1 is a dipeptide containing Ile, Val, Leu, Ac-Ile, Ac-Val, Ac-Leu, Gly-Ile, or Ac-Gly-Ile. Xaa2 and Xaa2 ＊ These are independently selected from Trp and its analogues, Xaa3 is His, Ala or an analogue of Ala, Phe, Trp or an analogue of Trp, Xaa4 is a dipeptide selected from L-Thr, D-Thr, Ile, Val, Gly, Thr-Ala, and Thr-Asn, or a tripeptide containing Thr-Ala-Asn, and any of the carboxyl-terminal -OH of L-Thr, D-Thr, Ile, Val, Gly, Al, or Asn may be -NH ２ A long-acting compstatin analog according to claim 90, which is replaced by the above.

92. The long-acting compstatin analog according to claim 90, wherein Xaa2 is an analog of Trp in which hydrophobicity is increased compared to Trp.

93. The long-acting compstatin analog according to claim 90, wherein Xaa2 is a substituted or unsubstituted bicyclic aromatic ring component or an analog of Trp comprising two or more substituted or unsubstituted monocyclic aromatic ring components.

94. Xaa2 ＊ The long-acting compstatin analog according to claim 90, wherein is an analog of Trp having an electronegative substituent on the indole ring and not having increased hydrophobicity compared to Trp.

95. Xaa2 ＊ The long-acting compstatin analog according to claim 90, wherein the compstatin analog is a Trp analog comprising a lower alkoxy substituent or a lower alkyl substituent at the 1st or 5th position of tryptophan, or a halogen substituent at the 5th or 6th position of tryptophan.

96. Xaa2 ＊ However, it is an analog of Trp containing a lower alkoxy substituent or lower alkyl substituent at the 1st or 5th position of tryptophan, or a halogen substituent at the 5th or 6th position of tryptophan, and Xaa2 ＊ A long-acting compstatin analog according to claim 90, wherein is Trp.

97. A long-acting compstatin analog according to claim 90, comprising a compstatin analog moiety containing a cyclic peptide having a sequence selected from the group consisting of SEQ ID NOs: 9 to 36, and optionally further comprising an amino acid having a side chain containing a primary, secondary, or sulfhydryl reactive group.

98. The long-acting compstatin analog according to claim 90, wherein the compstatin analog portion comprises a cyclic peptide having a sequence selected from the group consisting of SEQ ID NOs: 14, 21, 28, 29, 32, 33, 34, or 36.

99. The long-acting compstatin analog according to claim 90, wherein the compstatin analog portion comprises a cyclic peptide having the sequence of SEQ ID NO: 28, 32, or 34.

100. A long-acting compstatin analog according to any one of claims 54 to 83, having the sequence X'aa1-X'aa2-X'aa3-X'aa4-Gln-Asp-Xaa-Gly-X"aa1-X"aa2-X"aa3-X"aa4-X"aa5 (SEQ ID NO: 5), wherein the formula comprises a compstatin analog moiety containing a cyclic peptide selected from Trp and Trp analogs, where X'aa4 and Xaa are independently selected from amino acids and amino acid analogs, X'aa1, X'aa2, X'aa3, X"aa1, X"aa2, X"aa3, X"aa4 and X"aa5 are independently selected from amino acids and amino acid analogs, X'aa2 and X"aa4 are not Cys, and the peptide is cyclized via a bond between X'aa2 and X"aa4.

101. A long-acting compstatin analog according to claim 100, wherein one or more of X'aa1, X'aa3, X"aa2, X"aa3, and X"aa5 correspond to amino acids at the corresponding positions of the peptide according to any one of claims 90 to 99, and X"aa1 is Ala or a monomethyl unbranched amino acid.

102. A long-acting compstatin analog according to claim 100, wherein one of X'aa2 and X''aa4 is an amino acid or amino acid analog having a side chain containing a primary or secondary amine, and the other of X'aa2 and X''aa4 is an amino acid or amino acid analog having a side chain containing a carboxylic acid group, and the bond is an amide bond.

103. A long-acting compstatin analog according to claim 100, wherein X'aa1, X'aa3, X"aa1, X"aa2, X"aa3, and X"aa5 correspond to amino acids at the corresponding positions of the cyclic peptide according to any one of claims 90 to 99, and optionally one of X'aa2 and X"aa4 is an amino acid or amino acid analog having a side chain containing a primary or secondary amine, and the other of X'aa2 and X"aa4 is an amino acid or amino acid analog having a side chain containing a carboxylic acid group, and the bond is an amide bond.

104. A long-acting compstatin analog according to any one of claims 84 to 103, wherein the cyclic peptide is acetylated at the N-terminus, amidated at the C-terminus, or acetylated at the N-terminus and amidated at the C-terminus.

105. (i) A compound comprising a cyclic peptide having a sequence selected from the group consisting of SEQ ID NOs: 14, 21, 28, 29, 32, 33, 34, or 36; and (ii) A long-acting compstatin analog comprising a polymer having a molecular weight of 10 kD to 45 kD.

106. A long-acting compstatin analog according to any one of claims 84 to 105, comprising a compound obtained by reacting at least one NHS ester of any compound of formulas I to XVI or A to H with a side chain or terminal amino group of a compstatin analog moiety, wherein any compound of formulas I to XVI or A to H has a molecular weight of 10 kD to 45 kD.

107. A method for producing a long-acting compstatin analog, comprising reacting a compound of formulas I to XVI or A to H with a compstatin analog moiety, wherein the compound of formulas I to XVI or A to H has a molecular weight of 10 kD to 45 kD.

108. A method for producing a long-acting compstatin analog, comprising reacting a compound of formulas I to XVI or formulas A to H with a compstatin analog moiety containing any amino acid sequence of sequence numbers 3 to 36, 37, 37A, 38A, 39A, 40a, or 41A, wherein the compound of formulas I to XVI or formulas A to H has a molecular weight of 10 kD to 45 kD.

109. A long-acting compstatin analog prepared according to or having the same structure as described in any one of claims 107 to 108.

110. A long-acting compstatin analog according to any one of claims 84 to 106, 109, or 145 to 164, further comprising a targeting portion.

111. A composition comprising a long-acting compstatin analog according to any one of claims 54 to 105, 109, or 145 to 164 and a pharmaceutically acceptable carrier.

112. A pharmaceutical composition comprising a long-acting compstatin analog according to any one of claims 54 to 106, 109, or 145 to 164.

113. A pharmaceutical composition comprising a long-acting compstatin analog according to any one of claims 54 to 106, 109, or 145 to 164 and a pharmaceutically acceptable carrier.

114. A method for reducing the susceptibility of a cell or organ to complement-dependent injury, comprising contacting the cell with a long-acting compstatin analog or composition according to any one of claims 54-106, 109-113, or 144-165.

115. The method according to claim 114, wherein the cell or organ is a human cell or a human organ.

116. The method according to claim 114, wherein the cells are blood cells or the organ is the heart, kidney, liver, lung, or pancreas.

117. The method according to claim 114, comprising administering the long-acting compstatin analog or the composition to a subject.

118. A method for treating a subject requiring treatment for complement-mediated disorder, comprising administering to the subject a long-acting compstatin analog or composition according to any one of claims 54-106, 109-113, 144-165, or 167-174.

119. The method according to claim 118, comprising locally administering the long-acting compstatin analog to a site at risk of complement-mediated injury or a site where complement-mediated injury has occurred.

120. The method according to claim 118, wherein the disorder causes complement-mediated injury in red blood cells.

121. The method according to claim 118, wherein the disorder causes complement-mediated injury in red blood cells, and the long-acting compstatin analog is administered intravascularly or subcutaneously.

122. The method according to claim 118, wherein the subject has a defect in complement control.

123. The method according to claim 118, wherein the subject requires treatment for transplant rejection.

124. The method according to claim 118, wherein the subject requires treatment for ischemia / reperfusion injury.

125. The method according to claim 118, wherein the subject requires treatment for hemolytic anemia.

126. The method according to claim 118, wherein the subject requires treatment for an autoimmune disease.

127. The method according to claim 118, wherein the subject requires treatment for neuropathic pain.

128. The method according to claim 118, wherein the subject requires treatment with MPGN.

129. The method according to claim 118, wherein the subject requires treatment for neuromyelitis optica.

130. The method according to claim 118, wherein the subject requires treatment for spinal cord injury.

131. The method according to claim 118, wherein the subject requires treatment for asthma, COPD, or idiopathic pulmonary fibrosis.

132. The method according to any one of claims 118 to 131, comprising subcutaneously administering the long-acting compstatin analog.

133. The method according to any one of claims 118 to 131, comprising administering the long-acting compstatin analog subcutaneously once or more times a day.

134. The method according to any one of claims 118 to 131, comprising administering the long-acting compstatin analog transdermally.

135. The method according to any one of claims 118 to 131, wherein the long-acting compstatin analog is administered subcutaneously using a pen device.

136. The method according to any one of claims 118 to 131, wherein the long-acting compstatin analog is administered intramuscularly.

137. The method according to any one of claims 118 to 131, comprising a polymer having a molecular weight of approximately 40 kDa of a long-acting compstatin analog, administered subcutaneously once or twice daily, wherein the total daily dose is 90 mg to 360 mg, and optionally 180 mg / day to 270 mg / day.

138. The method according to claim 107 or 108, comprising a method in which any compound of formulas I to XVI or A to H and a compstatin analog moiety each undergo click functionality and a click chemical reaction.

139. Compstatin analogs containing a click chemical group.

140. The compstatin analog according to claim 139, wherein the compstatin analog comprises a compound containing any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A.

141. A compstatin analog according to claim 139 or 140, wherein the click chemical group comprises an azide, an alkyne, an octin, or a dibenzoarylcyclooctin.

142. The compstatin analog according to claim 140, wherein the click chemical group is DBCO, DIBO, DIFO, BARAC, or BCN.

143. The compstatin analog according to claim 140, wherein the click chemical group is suitable for copper-free click chemical reactions.

144. A composition comprising any compstatin analog of claims 139 to 143 and a CRM, wherein the CRM comprises a polymer having a molecular weight of 10 kD to 45 kD, optionally 35 kD to 45 kD, for example 40 kD.

145. A conjugate formed by the reaction of a compstatin analog of any of claims 139 to 143 with a CMR containing a complementary click chemical group, wherein the CRM comprises a polymer having a molecular weight of 10 kD to 45 kD, optionally 35 kD to 45 kD, for example 40 kD.

146. A long-acting compstatin analog comprising a compstatin analog moiety and a CRM linked via click chemical bonds, wherein the CRM comprises a polymer having a molecular weight of 10 kD to 45 kD, optionally 35 kD to 45 kD, for example 40 kD.

147. A long-acting compstatin analog comprising a compstatin analog moiety and a CRM, wherein the CRM comprises a POZ having a molecular weight of 10 kD to 45 kD, and optionally 35 kD to 45 kD, for example, 40 kD.

148. A long-acting compstatin analog comprising a CRM containing at least two compstatin analog moieties and a polymer having a molecular weight of 10 kD to 45 kD, optionally 35 kD to 45 kD, for example 40 kD, and optionally each compstatin analog moiety containing one of SEQ ID NOs: 3 to 36, 37, 37A, 38A, 39A, 40A, or 41A.

149. A long-acting compstatin analog according to claim 148, comprising a compstatin analog having the same sequence as the compstatin analog moiety, and having at least 90% activity or at least 100% activity on a molar basis.

150. A long-acting compstatin analog according to claim 148, wherein CRM contains PEG.

151. A long-acting compstatin analog according to claim 148, wherein CRM contains POZ.

152. The long-acting compstatin analog according to claim 148, wherein the CRM comprises a polypeptide.

153. A long-acting compstatin analog according to any one of claims 148 to 152, comprising two compstatin analog moieties.

154. A long-acting compstatin analog according to any one of claims 148 to 152, comprising three compstatin analog moieties.

155. A long-acting compstatin analog according to any one of claims 148 to 152, comprising 2 to 8 compstatin analog moieties.

156. The long-acting compstatin analog according to claim 148, comprising two compstatin analog moieties and a linear PEG, wherein the compstatin analog moieties are optionally bound to each end of the linear PEG by carbamate or ester bonds.

157. A long-acting compstatin analog according to claim 148, comprising three compstatin analog moieties and PEG.

158. A long-acting compstatin analog according to claim 148, comprising 2 to 8 compstatin analog moieties and PEG.

159. A long-acting compstatin analog according to claim 148, comprising two compstatin analog moieties and a POZ.

160. A long-acting compstatin analog according to claim 148, comprising three compstatin analog moieties and a POZ.

161. A long-acting compstatin analog according to claim 148, comprising 2 to 8 compstatin analog moieties and POZ.

162. A long-acting compstatin analog according to claim 148, comprising two compstatin analog moieties and a polypeptide.

163. A long-acting compstatin analog according to claim 148, comprising three compstatin analog moieties and a polypeptide.

164. A long-acting compstatin analog according to claim 148, comprising 2 to 8 compstatin analog moieties and polypeptides.

165. A composition comprising a compstatin analog according to any one of claims 139 to 164, which is optionally a pharmaceutical composition.

166. The method according to claim 118, wherein a long-acting compstatin analog is administered for the treatment of Th17-related disease.

167. A long-acting compstatin analog according to any one of claims 54-106, 109-113, 144-165, or 167-174, having a molecular weight of at least about 30 kD, a terminal phase half-life of at least about 3 days when administered to primates, and exhibiting at least 80% of the activity of a compstatin analog that contains the same compstatin analog sequence as the compstatin analog molar portion but is not bound to CRM.

168. A long-acting compstatin analog according to claim 167, having a molecular weight of at least about 40 kD.

169. A long-acting compstatin analog according to claim 167 or 168, having a terminal phase half-life of at least about four days.

170. A long-acting compstatin analog according to claim 167 or 168, having a terminal phase half-life of at least about 5 days.

171. A long-acting compstatin analog according to claim 167 or 168, which, on a molar basis, contains the same compstatin analog sequence as the compstatin analog moiety but exhibits at least 90% of the activity of a compstatin analog that is not bound to CRM.

172. A long-acting compstatin analog according to claim 167 or 168, which, on a molar basis, contains the same compstatin analog sequence as the compstatin analog moiety but has at least equivalent activity to a compstatin analog that is not bound to CRM.

173. A long-acting compstatin analog according to claim 167 or 168, wherein the compstatin analog portion comprises any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A.

174. A long-acting compstatin analog according to any one of claims 167 to 173, wherein the CRM comprises PEG, POZ, or polypeptide.

175. A method for treating complement-mediated ocular disorders, comprising administering a long-acting compstatin analog described in any of claims 54-106, 109-113, 144-165, or 167-174 to a subject requiring treatment.

176. The method according to claim 175, wherein the eye disorder is AMD.

177. The method according to claim 175, wherein the ocular disorder is geographic atrophy.

178. The method according to claim 175, wherein the eye disorder is intermediate-type AMD.

179. The method according to any one of claims 175 to 178, wherein a long-acting compstatin analog is administered by intravitreal injection.

180. The method according to claim 179, wherein LACA is administered monthly or every other month.

181. The method according to claim 179 or 180, wherein the dose administered is 10 mg to 20 mg.

182. The method according to claim 179 or 180, wherein the dose administered is 15 mg.

183. A unit dose of LACA containing a polymer having a molecular weight of 10 kD to 45 kD, and optionally 35 kD to 45 kD, for example 40 kD, wherein the amount of the unit dose is 45 mg to 360 mg, and optionally 180 mg to 270 mg, for example 180 mg or 270 mg.

184. A unit dose for intravitreal administration of LACA, wherein LACA comprises a polymer having a molecular weight of 10 kD to 45 kD, optionally 35 kD to 45 kD, for example 40 kD, and the amount of the unit dose is 10 mg to 20 mg, optionally 15 mg.

185. The unit dose according to claim 183 or 184, wherein the polymer has a molecular weight of about 30 kD.

186. The unit dose according to claim 183 or 184, wherein the polymer has a molecular weight of about 40 kD.

187. The unit dose according to any one of claims 183 to 186, further comprising a pharmaceutically acceptable carrier.

188. A syringe or container comprising the unit dose according to any one of claims 183 to 187.

189. The unit dose, syringe, or container according to claim 187 or 188, wherein LACA is present in a pharmaceutically acceptable carrier at a concentration of 125 mg / ml to 200 mg / ml.

190. The unit dose, syringe, or container according to claim 189, wherein LACA is present in a pharmaceutically acceptable carrier at a concentration of 140 mg / ml to 180 mg / ml, for example, 150 mg / ml.

191. The unit dose, syringe, or container according to claim 187 or 188, wherein LACA is present in a pharmaceutically acceptable carrier at a concentration of 80 mg / ml to 125 mg / ml, for example, 100 mg / ml.

192. A unit dose, syringe, or container according to any one of claims 183 to 191, wherein the polymer is PEG.

193. A unit dose, syringe, or container according to any one of claims 183 to 192, wherein LACA comprises two compstatin analog moieties, where one compstatin analog moiety is bonded to each end of the polymer by a moiety comprising an unsaturated alkyl moiety, a moiety comprising a non-aromatic cyclic ring system, an aromatic moiety, an ether moiety, an amide moiety, an ester moiety, a carbonyl moiety, an imine moiety, a thioether moiety, and / or an amino acid residue.

194. A unit dose, syringe, or container according to any one of claims 183 to 193, wherein LACA comprises two compstatin analog moieties, where one compstatin analog moiety is bonded to each end of the polymer by an ester or carbamate bond.

195. A unit dose, syringe, or container according to any one of claims 183 to 194, wherein the compstatin analog portion comprises any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A.

196. A unit dose, syringe, or container according to any one of claims 183 to 195, wherein the compstatin analog portion comprises a peptide containing a cyclic portion, the peptide being extended at its N-terminus, C-terminus, or both ends by one or more amino acids, where at least one of the amino acids has a side chain containing a reactive ring, and where the extension of one or more amino acids is optionally separated from the cyclic portion of the compstatin analog portion by a spacer.

197. The unit dose, syringe, or container according to claim 196, wherein the spacer comprises a substituted or unsubstituted, saturated or unsaturated alkyl chain, or oligo(ethylene glycol) chain.

198. The unit dose, syringe, or container according to claim 197, wherein the spacer includes an AEEAc portion.

199. A unit dose, syringe, or container according to any one of claims 183 to 198, wherein LACA is CA28-2TS-BF or CA28-2GS-BF.

200. A unit dose of LACA containing a polymer having a molecular weight of 35 kD to 45 kD, wherein the unit dose is 545 mg to 5040 mg.

201. The unit dose according to claim 200, wherein the polymer has a molecular weight of 37.5 kD to 42.5 kD.

202. The unit dose according to claim 200 or 201, wherein the polymer has a molecular weight of 40 kD.

203. The unit dose according to any one of claims 200 to 202, wherein the amount is 545 mg to 1690 mg, and in some cases, the unit dose is for subcutaneous administration three times a week.

204. The unit dose according to any one of claims 200 to 202, wherein the amount is 630 mg to 930 mg, and in some cases, the unit dose is for subcutaneous administration three times a week.

205. The unit dose according to any one of claims 200 to 202, wherein the amount is 795 mg to 885 mg, and in some cases, the unit dose is for subcutaneous administration three times a week.

206. The unit dose according to any one of claims 200 to 202, wherein the amount is 585 mg to 2510 mg, and in some cases the unit dose is for subcutaneous administration twice a week.

207. The unit dose according to any one of claims 200 to 202, wherein the amount is 900 mg to 1395 mg, and in some cases, the unit dose is for subcutaneous administration twice a week.

208. The unit dose according to any one of claims 200 to 202, wherein the amount is 990 mg to 1215 mg, and in some cases, the unit dose is for subcutaneous administration twice a week.

209. The unit dose according to any one of claims 200 to 202, wherein the amount is 1215 mg to 1395 mg, and in some cases, the unit dose is for subcutaneous administration twice a week.

210. The unit dose according to any one of claims 200 to 202, wherein the amount is 1080 mg to 5040 mg, and in some cases, the unit dose is for subcutaneous administration once a week.

211. The unit dose according to any one of claims 200 to 202, wherein the amount is 2160 mg to 2520 mg, and in some cases, the unit dose is for subcutaneous administration once a week.

212. The unit dose according to any one of claims 200 to 202, wherein the amount is 2520 mg to 2880 mg, and in some cases, the unit dose is for subcutaneous administration once a week.

213. The unit dose according to any one of claims 200 to 202, wherein the amount is 2880 mg to 3240 mg, and in some cases, the unit dose is for subcutaneous administration once a week.

214. The unit dose according to any one of claims 200 to 202, wherein the amount is 3240 mg to 3600 mg, and in some cases, the unit dose is for once-weekly subcutaneous administration.

215. A unit dose according to any one of claims 200 to 214, further comprising a pharmaceutically acceptable carrier.

216. A syringe or container containing the unit dose according to any one of claims 200 to 216.

217. A unit dose, syringe, or container according to any one of claims 200 to 216, wherein LACA is present in a pharmaceutically acceptable carrier at a concentration of 25 mg / ml to 150 mg / ml.

218. The unit dose, syringe, or container according to claim 217, wherein LACA is present in a pharmaceutically acceptable carrier at a concentration of 25 mg / ml to 50 mg / ml.

219. The unit dose, syringe, or container according to claim 217, wherein LACA is present in a pharmaceutically acceptable carrier at a concentration of 50 mg / ml to 75 mg / ml.

220. The unit dose, syringe, or container according to claim 217, wherein LACA is present in a pharmaceutically acceptable carrier at a concentration of 75 mg / ml to 100 mg / ml.

221. The unit dose, syringe, or container according to claim 217, wherein LACA is present in a pharmaceutically acceptable carrier at a concentration of 100 mg / ml to 125 mg / ml.

222. A unit dose, syringe, or container according to any one of claims 200 to 221, wherein the polymer is PEG.

223. A unit dose, syringe, or container according to any one of claims 200 to 222, wherein the polymer is a linear polymer, and LACA comprises two compstatin analog moieties, each of which is bonded to the ends of the polymer by a moiety containing an unsaturated alkyl moiety, a moiety containing a non-aromatic ring system, an aromatic moiety, an ether moiety, an amide moiety, an ester moiety, a carbonyl moiety, an imine moiety, a thioether moiety, and / or an amino acid residue.

224. The unit dose, syringe, or container according to claim 223, wherein one compstatin analog moiety is attached to each end of the polymer by an ester or carbamate bond.

225. The unit dose, syringe, or container according to claim 223 or 224, wherein LACA comprises two compstatin analog moieties, one of which is attached to each end of the polymer by a carbamate bond.

226. A unit dose, syringe, or container according to any one of claims 200 to 225, wherein the compstatin analog portion comprises any of SEQ ID NOs: 3-36, 37, 37A, 38A, 39A, 40A, or 41A.

227. The unit dose, syringe, or container according to any one of claims 200 to 226, wherein the compstatin analog portion comprises a peptide elongated by one or more amino acids at the N-terminus, C-terminus, or both ends, including a cyclic portion, and the elongation of one or more amino acids may be separated from the cyclic portion of the compstatin analog portion by a spacer, as is sometimes the case.

228. The unit dose, syringe, or container according to claim 227, wherein the cyclic peptide is extended by an amino acid sequence comprising at least one amino acid having a side chain containing a primary or secondary amine.

229. A unit dose, syringe, or container according to claim 227 or 228, wherein the lysine is a cyclic peptide in which at least one amino acid having a side chain containing a primary or secondary amine is linked at the C-terminus.

230. A unit dose, syringe, or container according to any one of claims 227 to 229, wherein the spacer comprises a substituted or unsubstituted, saturated or unsaturated alkyl chain, or an oligo(ethylene glycol) chain.

231. The oligo(ethylene glycol) portion is (-(O-CH2-CH2-) ｎ The unit dose, syringe or container according to claim 230, wherein n is 1 to 10.

232. The spacer is a covalently bonded -(CH2) where m is 1 to 10 and n is 1 to 10. ｍ - and - (O-CH2-CH2-) ｎ The method according to any one of claims 227 to 231, including the method described in any one of claims 227 to 231.

233. The unit dose, syringe, or container according to claim 232, wherein the spacer comprises 8-amino-3,6-dioxaoctanoic acid (AEEAc) or 11-amino-3,6,9-trioxaundecanoic acid.

234. The unit dose, syringe, or container according to claim 233, wherein the spacer includes an AEEAc portion.

235. A unit dose, syringe, or container according to any one of claims 200 to 234, wherein LACA is CA28-2TS-BF or CA28-2GS-BF.

236. A method for inhibiting complement activation in a subject, comprising subcutaneously administering a unit dose according to any one of claims 200 to 235.

237. The method according to claim 236, wherein a unit dose is administered using a syringe pump.

238. The method according to claim 236, wherein a unit dose is administered using an intracellular delivery device.

239. The method according to any one of claims 236 to 238, wherein the unit dose is administered in a volume of 10 ml to 50 ml.

240. The method according to any one of claims 236 to 239, wherein the unit dose is administered in a volume of 20 ml to 40 ml.

241. The method according to any one of claims 236 to 240, wherein a unit dose is administered three times a week.

242. The method according to any one of claims 236 to 240, wherein a unit dose is administered twice a week.

243. The method according to any one of claims 236 to 240, wherein a unit dose is administered once a week.

244. The method according to any one of claims 236 to 243, wherein the subject is suffering from or at risk of complement-mediated disorder.

245. A method for treating a subject requiring treatment for complement-mediated disorder, comprising subcutaneously administering a unit dose according to any one of claims 200 to 235 to the subject.

246. The method according to claim 245, wherein a unit dose is administered using a syringe pump.

247. The method according to claim 245, wherein a unit dose is administered using an intracellular delivery device.

248. The method according to any one of claims 245 to 247, wherein the unit dose is administered in a volume of 10 ml to 50 ml.

249. The method according to any one of claims 245 to 247, wherein the unit dose is administered in a volume of 20 ml to 40 ml, and optionally in a volume of 20 ml, 21 ml, 22 ml, 23 ml, 24 ml, or 25 ml.

250. The method according to any one of claims 245 to 249, wherein a unit dose is administered three times a week.

251. The method according to any one of claims 245 to 249, wherein a unit dose is administered twice a week.

252. The method according to any one of claims 245 to 249, wherein a unit dose is administered once a week.

253. The method according to any one of claims 244 to 252, wherein the complement-mediated disorder is hemolytic anemia.

254. The method according to any one of claims 244 to 252, wherein the complement-mediated disorder is PNH.

255. The method according to any one of claims 244 to 252, wherein the complement-mediated disorder is autoimmune hemolytic anemia, and in some cases the hemolytic anemia is cold agglutinin disease or warm autoimmune hemolytic anemia.

256. The method according to any one of claims 244 to 252, wherein the complement-mediated disorder is myasthenia gravis.

257. The method according to any one of claims 244 to 252, wherein the complement-mediated disorder is NMO.

258. The method according to any one of claims 244 to 252, wherein the complement-mediated disorder is polyneuropathy, or the complement-mediated disorder is neuropathy, or the complement-mediated disorder is vasculitis.

259. A method for inhibiting complement activation in a subject, comprising subcutaneously administering LACA containing a polymer having a molecular weight of 35 kD to 45 kD to a subject, according to a dosing schedule of administering LACA three times a week, twice a week, or once a week.

260. The method according to claim 259, wherein the polymer has a molecular weight of 37.5 kD to 42.5 kD.

261. The method according to claim 259 or 260, wherein the polymer has a molecular weight of 40 kD.

262. The method according to any one of claims 259 to 261, wherein LACA is administered three times a week in an amount of 545 mg to 1690 mg per dose.

263. The method according to any one of claims 259 to 261, wherein LACA is administered three times a week in an amount of 630 mg to 930 mg per dose.

264. The method according to any one of claims 259 to 261, wherein LACA is administered three times a week in an amount of 795 mg to 885 mg per dose.

265. The method according to any one of claims 259 to 261, wherein LACA is administered twice a week in an amount of 585 mg to 2510 mg per dose.

266. The method according to any one of claims 259 to 261, wherein LACA is administered twice a week in an amount of 900 mg to 1395 mg per dose.

267. The method according to any one of claims 259 to 261, wherein LACA is administered twice a week in an amount of 990 mg to 1215 mg per dose.

268. The method according to any one of claims 259 to 261, wherein LACA is administered twice a week in an amount of 1215 mg to 1395 mg per dose.

269. The method according to any one of claims 259 to 261, wherein LACA is administered once a week in an amount of 1080 mg to 5040 mg per dose.

270. The method according to any one of claims 259 to 261, wherein LACA is administered once a week in an amount of 2160 mg to 2520 mg per dose.

271. The method according to any one of claims 259 to 261, wherein LACA is administered once a week in an amount of 2520 mg to 2880 mg per dose.

272. The method according to any one of claims 259 to 261, wherein LACA is administered once a week in an amount of 2880 mg to 3240 mg per dose.

273. The method according to any one of claims 259 to 261, wherein LACA is administered once a week in an amount of 3240 mg to 3600 mg per dose.

274. The method according to any one of claims 259 to 261, wherein LACA is administered as a composition comprising LACA and a pharmaceutically acceptable carrier.

275. The method according to any one of claims 259 to 274, wherein LACA is administered at a concentration of 25 mg / ml to 150 mg / ml.

276. The method according to any one of claims 259 to 274, wherein LACA is administered at a concentration of 25 mg / ml to 50 mg / ml, and optionally at a concentration of 45 mg / ml to 50 mg / ml.

277. The method according to any one of claims 259 to 274, wherein LACA is administered at a concentration of 50 mg / ml to 75 mg / ml, optionally at a concentration of 50 mg / ml to 60 mg / ml, and optionally at a concentration of 50 mg / ml, 51 mg / ml, 52 mg / ml, 53 mg / ml, 54 mg / ml, 55 mg / ml, 56 mg / ml, 57 mg / ml, 58 mg / ml, 59 mg / ml, or 60 mg / ml.

278. The method according to any one of claims 259 to 274, wherein LACA is administered at a concentration of 75 mg / ml to 100 mg / ml.

279. The method according to any one of claims 259 to 274, wherein LACA is administered at a concentration of 100 mg / ml to 125 mg / ml.

280. The method according to any one of claims 259 to 279, wherein the polymer is PEG.

281. The method according to any one of claims 259 to 280, wherein the polymer is a linear polymer, and LACA comprises two compstatin analog moies, each of which is bonded to the ends of the polymer by a moiety containing an unsaturated alkyl moiety, a moiety containing a non-aromatic ring system, an aromatic moiety, an ether moiety, an amide moiety, an ester moiety, a carbonyl moiety, an imine moiety, a thioether moiety and / or an amino acid residue.

282. The method according to any one of claims 259 to 281, wherein LACA comprises two compstatin analog moieties, one of which is attached to each end of the polymer by an ester or carbamate bond.

283. The method according to any one of claims 259 to 282, wherein LACA comprises two compstatin analog moieties, one of which is attached to each end of the polymer by a carbamate bond.

284. The method according to any one of claims 259 to 283, wherein the compstatin analog portion comprises any of SEQ ID NOs: 3 to 36, 37, 37A, 38A, 39A, 40A, or 41A.

285. The method according to any one of claims 259 to 284, wherein the compstatin analog moiety comprises a peptide elongated by one or more amino acids at the N-terminus, C-terminus, or both ends, including a cyclic moiety, and the elongation of one or more amino acids may be separated from the cyclic moiety of the compstatin analog moiety by a spacer.

286. A unit dose, syringe, or container according to claim 285, which is extended by an amino acid sequence comprising at least one amino acid having a side chain containing a primary or secondary amine.

287. The method according to claim 285 or 286, wherein the lysine is a cyclic peptide to which at least one amino acid having a side chain containing a primary or secondary amine is linked at the C-terminus.

288. The method according to any one of claims 285 to 287, wherein the spacer comprises a substituted or unsubstituted, saturated or unsaturated alkyl chain, or an oligo(ethylene glycol) chain.

289. The oligo(ethylene glycol) portion is (-(O-CH ２ -CH ２ -) ｎ The method according to claim 288, wherein n is 1 to 10.

290. The spacer is a covalently bonded -(CH2) where m is 1 to 10 and n is 1 to 10. ｍ - and - (O-CH2-CH2-) ｎ The method according to any one of claims 285 to 289, including the method described in any one of claims 285 to 289.

291. The method according to claim 290, wherein the spacer comprises 8-amino-3,6-dioxaoctanoic acid (AEEAc) or 11-amino-3,6,9-trioxaundecanoic acid.

292. The method according to claim 291, wherein the spacer includes an AEEAc portion.

293. The method according to any one of claims 259 to 292, wherein LACA is CA28-2TS-BF or CA28-2GS-BF.

294. The method according to any one of claims 259 to 293, wherein the subject is suffering from complement-mediated disorder.

295. A method for treating subjects requiring treatment of complement-mediated disorders, comprising subcutaneous administration of LACA containing polymers having a molecular weight of 35 kD to 45 kD, according to a dosing schedule of administering LACA three times a week, twice a week, or once a week.

296. The method according to claim 295, wherein the polymer has a molecular weight of 37.5 kD to 42.5 kD.

297. The method according to claim 295 or 296, wherein the polymer has a molecular weight of 40 kD.

298. The method according to any one of claims 295 to 297, wherein LACA is administered three times a week in an amount of 545 mg to 1690 mg per dose.

299. The method according to any one of claims 295 to 297, wherein LACA is administered three times a week in an amount of 630 mg to 930 mg per dose.

300. The method according to any one of claims 295 to 297, wherein LACA is administered three times a week in an amount of 795 mg to 885 mg per dose.

301. The method according to any one of claims 295 to 297, wherein LACA is administered twice a week in an amount of 585 mg to 2510 mg per dose.

302. The method according to any one of claims 295 to 297, wherein LACA is administered twice a week in an amount of 900 mg to 1395 mg per dose.

303. The method according to any one of claims 295 to 297, wherein LACA is administered twice a week in an amount of 990 mg to 1215 mg per dose, and optionally in an amount of at least 1044 mg, 1056 mg, 1060 mg, 1078 mg, 1080 mg, 1100 mg, 1104 mg, 1120 mg, 1122 mg, 1125 mg, 1140 mg, 1144 mg, 1150 mg, or 1170 mg.

304. The method according to any one of claims 295 to 297, wherein LACA is administered twice a week in an amount of 1215 mg to 1395 mg per dose.

305. The method according to any one of claims 295 to 297, wherein LACA is administered once a week in an amount of 1080 mg to 5040 mg per dose.

306. The method according to any one of claims 295 to 297, wherein LACA is administered once a week in an amount of 2160 mg to 2520 mg per dose.

307. The method according to any one of claims 295 to 297, wherein LACA is administered once a week in an amount of 2520 mg to 2880 mg per dose.

308. The method according to any one of claims 295 to 297, wherein LACA is administered once a week in an amount of 2880 mg to 3240 mg per dose.

309. The method according to any one of claims 295 to 297, wherein LACA is administered once a week in an amount of 3240 mg to 3600 mg per dose.

310. The method according to any one of claims 295 to 297, wherein LACA is administered as a composition comprising LACA and a pharmaceutically acceptable carrier.

311. The method according to any one of claims 295 to 310, wherein LACA is administered at a concentration of 25 mg / ml to 150 mg / ml.

312. LACA is administered at a concentration of 25 mg / ml to 50 mg / ml, and optionally (a) at a concentration of 45 mg / ml, 46 mg / ml, 47 mg / ml, 48 mg / ml, 49 mg / ml or 50 mg / ml; (b) the LACA is administered twice a week in amounts of 990 mg to 1215 mg per dose, and optionally 1056 mg to 1200 mg per dose, for example 1056 mg, 1078 mg, 1080 mg, 1100 mg, 1104 mg, 1125 mg, 1140 mg, 1150 mg or 1170 mg per dose; (c) each dose is administered in a volume of 22 ml to 25 ml, with a concentration of LACA of 45 mg / ml to 50 mg / ml; (d) LACA at 49 mg / ml The method according to any one of claims 295 to 310, wherein (e) 22 ml of the composition containing is administered via SC twice a week; (f) 23 ml of the composition containing 48 mg / ml of LACA is administered via SC twice a week; (g) 24 ml of the composition containing 46 mg / ml of LACA is administered via SC twice a week; (h) 25 ml of the composition containing 44 mg / ml of LACA is administered via SC twice a week; and / or (i) 25 ml of the composition containing 45 mg / ml of LACA is administered via SC twice a week; (j) 25 ml of the composition containing 46 mg / ml of LACA is administered via SC twice a week; and / or (k) 26 ml of the composition containing 45 mg / ml of LACA is administered via SC twice a week.

313. LACA is administered at a concentration of 50 mg / ml to 75 mg / ml, and depending on the case, (a) the concentration may be 50 mg / ml, 51 mg / ml, 52 mg / ml, 53 mg / ml, 54 mg / ml, 55 mg / ml, 56 mg / ml, 57 mg / ml, 58 mg / ml, 59 mg / ml, or 60 mg / ml; or (b) the LACA is administered twice a week in doses of 990 mg to 1215 mg. (c) Each dose is administered in an amount of 1044 mg to 1200 mg, for example, 1044 mg, 1060 mg, 1080 mg, 1120 mg, 1122 mg, 1125 mg, 1140 mg, or 1144 mg per dose; (d) Each dose is administered in a volume of 18 ml to 25 ml, with a concentration of LACA of 50 mg / ml to 60 mg / ml; (d) 58 mg / m³ The method according to any one of claims 295 to 310, comprising: (i) administering 18 ml of a composition containing LACA at a dose of l twice a week via SC; (e) administering 18 ml of a composition containing 60 mg / ml of LACA at a dose of SC twice a week; (f) administering 18 ml of a composition containing 60 mg / ml of LACA at a dose of SC twice a week; (g) administering 20 ml of a composition containing 54 mg / ml of LACA at a dose of SC twice a week; (h) administering 20 ml of a composition containing 55 mg / ml of LACA at a dose of SC twice a week; (i) administering 20 ml of a composition containing 56 mg / ml of LACA at a dose of SC twice a week; (j) administering 22 ml of a composition containing 50 mg / ml of LACA at a dose of SC twice a week; (k) administering 22 ml of a composition containing 51 mg / ml of LACA at a dose of SC twice a week; and (l) administering 22 ml of a composition containing 52 mg / ml of LACA at a dose of SC twice a week.

314. The method according to any one of claims 295 to 310, wherein LACA is administered at a concentration of 75 mg / ml to 100 mg / ml.

315. The method according to any one of claims 295 to 310, wherein LACA is administered at a concentration of 100 mg / ml to 125 mg / ml.

316. The method according to any one of claims 295 to 315, wherein the polymer is PEG.

317. The method according to any one of claims 295 to 316, wherein the polymer is a linear polymer, and LACA comprises two compstatin analog moies, each of which is bonded to the ends of the polymer by an unsaturated alkyl moiety, a moiety containing a non-aromatic ring system, an aromatic moiety, an ether moiety, an amide moiety, an ester moiety, a carbonyl moiety, an imine moiety, a thioether moiety and / or an amino acid residue.

318. The method according to any one of claims 295 to 317, wherein LACA comprises two compstatin analog moieties, one of which is attached to each end of the polymer by an ester or carbamate bond.

319. The method according to any one of claims 295 to 318, wherein LACA comprises two compstatin analog moieties, one of which is attached to each end of the polymer by a carbamate bond.

320. The method according to any one of claims 295 to 319, wherein the compstatin analog portion comprises any of SEQ ID NOs: 3 to 36, 37, 37A, 38A, 39A, 40A, or 41A.

321. The method according to any one of claims 295 to 320, wherein the compstatin analog moiety comprises a peptide elongated by one or more amino acids at the N-terminus, C-terminus, or both ends, including a cyclic moiety, and the elongation of one or more amino acids may be separated from the cyclic moiety of the compstatin analog moiety by a spacer.

322. A unit dose, syringe, or container according to claim 321, which is extended by an amino acid sequence comprising at least one amino acid having a side chain containing a primary or secondary amine.

323. The method according to claim 321 or 322, wherein the lysine is a cyclic peptide to which at least one amino acid having a side chain containing a primary or secondary amine is linked at the C-terminus.

324. The method according to any one of claims 321 to 333, wherein the spacer comprises a substituted or unsubstituted, saturated or unsaturated alkyl chain, or an oligo(ethylene glycol) chain.

325. The oligo(ethylene glycol) portion is (-(O-CH2-CH2-) ｎ The method according to claim 324, wherein n is 1 to 10.

326. The spacer is a covalently bonded -(CH2) where m is 1 to 10 and n is 1 to 10. ｍ - and - (O-CH2-CH2-) ｎ The method according to any one of claims 321 to 325, including the method described in any one of claims 321 to 325.

327. The method according to claim 326, wherein the spacer comprises 8-amino-3,6-dioxaoctanoic acid (AEEAc) or 11-amino-3,6,9-trioxaundecanoic acid.

328. The method according to claim 327, wherein the spacer includes an AEEAc portion.

329. The method according to any one of claims 295 to 328, wherein LACA is CA28-2TS-BF or CA28-2GS-BF.

330. The method according to any one of claims 294 to 329, wherein the complement-mediated disorder is hemolytic anemia.

331. The method according to any one of claims 294 to 329, wherein the complement-mediated disorder is PNH.

332. The method according to any one of claims 294 to 329, wherein the complement-mediated disorder is autoimmune hemolytic anemia, and in some cases the hemolytic anemia is cold agglutinin disease or warm autoimmune hemolytic anemia.

333. The method according to any one of claims 294 to 329, wherein the complement-mediated disorder is myasthenia gravis.

334. The method according to any one of claims 294 to 329, wherein the complement-mediated disorder is NMO.

335. The method according to any one of claims 294 to 329, wherein the complement-mediated disorder is polyneuropathy, or the complement-mediated disorder is neuropathy, or the complement-mediated disorder is vasculitis.

336. A unit dose, syringe, container, or method according to any one of claims 183 to 335, wherein the compstatin analog comprises SEQ ID NO:

28.

337. A unit dose, syringe, container, or method according to any one of claims 175 to 336, wherein a long-acting compstatin analog is present in a composition comprising an excipient selected from the group consisting of one or more sugar alcohols and non-reducing sugars.

338. The unit dose, syringe, container, or method according to claim 337, wherein the excipient is trehalose or sorbitol.

339. The unit dose, syringe, container, or method according to claim 337 or 338, wherein the composition has a pH of 4.8 to 5.

2.

340. The unit dose, syringe, container, or method according to any one of claims 337 to 340, wherein the composition comprises sodium acetate as a buffering agent.

341. A unit dose, syringe, container, or method according to any one of claims 337 to 340, wherein the composition has an osmotic pressure of 250 mOsm to 380 mOsm, and optionally, an osmotic pressure of 280 mOsm to 350 mOsm.

342. A composition comprising approximately 150 mg / ml of CA28-2GS-BF or CA28-2TS-BF and a 0.50% to 0.55% NaCl aqueous solution.

343. The composition according to claim 342, wherein the concentration of NaCl is 0.51% to 0.54%, and optionally 0.52% to 0.53%.

344. A composition containing approximately 150 mg / ml of CA28-2GS-BF or CA28-2TS-BF, and 3.0% to 3.6% of sorbitol.

345. The composition according to claim 344, wherein the concentration of sorbitol is 3.1%, 3.2%, 3.3%, 3.4%, or 3.5%.

346. A composition comprising approximately 150 mg / ml of CA28-2GS-BF or CA28-2TS-BF and trehalose.

347. The composition according to claim 346, wherein the concentration of trehalose is 6.6% to 7.0%, and optionally 6.7%, 6.8%, or 6.9%.

348. The composition according to any one of claims 342 to 347, wherein the concentration of CA28-2GS-BF or CA28-2TS-BF is 145 mg / ml to 155 mg / ml, and optionally 150 mg / ml.

349. A composition according to any one of claims 342 to 348, comprising approximately 20 mM sodium acetate.

350. A composition according to any one of claims 342 to 349, having a volume of approximately 100 μL.

351. A composition according to any one of claims 342 to 349 for intravitreal administration.

352. A composition for use in the treatment of an eye disorder, wherein the eye disorder is, optionally, age-related macular degeneration, according to any one of claims 342 to 351.

353. The composition according to claim 352, wherein the eye disorder is AMD characterized by geographic atrophy.

354. A composition comprising CA28-2GS-BF or CA28-2TS-BF and sorbitol.

355. The composition according to claim 354, wherein the concentration of CA28-2GS-BF or CA28-2TS-BF is approximately 55 mg / ml, and optionally 52 mg / ml to 57 mg / ml.

356. The composition according to claim 355, wherein the concentration of CA28-2GS-BF or CA28-2TS-BF is 53 mg / ml, 54 mg / ml, or 55 mg / ml.

357. A composition according to any one of claims 354 to 356 for subcutaneous administration.

358. A composition according to any one of claims 354 to 357 for use in the treatment of complement-mediated disorders.

359. The composition according to claim 358, wherein the disorder is complement-mediated damage to red blood cells, and the disorder may be PNH or AIHA.

360. The composition according to claim 359, wherein the disorder is neuromyelitis optica, myasthenia gravis, or nephropathy.

361. The composition according to any one of claims 342 to 360, wherein the pH of the composition is 4.8 to 5.2, and optionally 5.0.

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