WO2024025440A1

WO2024025440A1 - Pharmaceutical composition of anti-cd20 antibody and use thereof

Info

Publication number: WO2024025440A1
Application number: PCT/RU2023/050174
Authority: WO
Inventors: Aleksandr Olegovich IAKOVLEV; Ekaterina Aleksandrovna LOMKOVA; Aleksandra Aleksandrovna SOZONOVA; Arina Valerevna ZINKINA-ORIKHAN; Daria Dmitrievna BOLSUN; Dmitry Valentinovich MOROZOV
Original assignee: Joint Stock Company "Biocad"
Priority date: 2022-07-26
Filing date: 2023-07-17
Publication date: 2024-02-01
Also published as: TW202417044A

Abstract

The present invention relates to the field of pharmacy and medicine, specifically to pharmaceutical compositions of anti-CD20 antibody, in particular divozilimab, which compositions may be used to treat a CD20-mediated disease or disorder. The invention further relates to the use of said compositions for the treatment of a CD20-mediated disease or disorder, as well as to a method of treating a CD20-mediated disease or disorder, comprising administering said compositions.

Description

PHARMACEUTICAL COMPOSITION OF ANTI-CD20 ANTIBODY AND USE THEREOF

Field of the invention

The present invention relates to the field of pharmacy and medicine, specifically to pharmaceutical compositions of anti-CD20 antibody, in particular divozilimab, which compositions may be used to treat a CD20-mediated disease or disorder.

Background of the invention

Lymphocytes are one of several populations of white blood cells; they specifically recognize and respond to a foreign antigen. The three major classes of lymphocytes are B lymphocytes (B cells), T lymphocytes (T cells) and natural killer (NK) cells. B lymphocytes are the cells responsible for antibody production and humoral immune response. B cells mature within the bone marrow and leave the marrow expressing an antigen-binding antibody on their cell surface. When a naive B cell first encounters the antigen for which the membrane-bound antibody is specific, the cell begins to divide rapidly and its progeny differentiate into memory B cells and effector cells called "plasma cells". The memory B cells have a long life span and they continue to express membrane-bound antibody with the same specificity as the original parent B cell. Plasma cells do not produce membrane bound antibody, instead they produce a secreted form of the antibody. Secreted antibodies are the major effector molecules of the humoral immune response.

The CD20 antigen (also called human B -lymphocyte-restricted differentiation antigen, Bp35) is a hydrophobic transmembrane protein with a molecular weight of approximately 35 kDa located on pre-B and mature B lymphocytes (Valentine et al. J. Biol. Chem. 264(19): 1989, pp.l 1282-11287; and Einfeld et al. EMBO J. 7(3), 1988, pp. 711-717). The antigen is also expressed on greater than 90% of B cell non-Hodgkin's lymphomas (NHL) (Anderson et al. Blood 63(6), 1984, pp. 1424-1433), but is not found on hematopoietic stem cells, pro-B cells, normal plasma cells or other normal tissues (Tedder et al., J. Immunol. 135(2), 1985, pp. 973-979). CD20 is thought to regulate an early step(s) in the activation process for cell cycle initiation and differentiation (Tedder et al., supra) and possibly functions as calcium ion channels (Tedder et al., J. Cell. Biochem. 14D, 1990, p. 195).

The prior art provides anti-CD20 antibodies: rituximab, obinutuzumab, ofatumumab, ocrelizumab and others.

Also, the prior art provides pharmaceutical compositions of various anti-CD20 antibodies. Known is a pharmaceutical composition comprising rituximab, sodium citrate, polysorbate 80, sodium chloride, hydrochloric acid and sodium hydroxide (Mabthera, concentrate for solution for infusion, Russian registration certificate number N013127/01). Known is a pharmaceutical composition comprising rituximab, recombinant human hyaluronidase, histidine, histidine hydrochloride monohydrate, trehalose dihydrate, methionine, polysorbate 80 (Mabthera, solution for subcutaneous administration, Russian registration certificate number (000358)-(PT[RG]-RU)). Known is a pharmaceutical composition comprising obinutuzumab, histidine, histidine hydrochloride monohydrate, trehalose dihydrate, pol oxamer 188 (Gazyva, concentrate for solution for infusion, Russian registration certificate number JIH[LP]-002867). Known is a pharmaceutical composition comprising ofatumumab, arginine, sodium acetate trihydrate, sodium chloride, polysorbate 80, EDTA disodium salt dihydrate, hydrochloric acid (Kesimpta, EMEA/H/C/005410). Known is a pharmaceutical composition comprising ocrelizumab, sodium acetate trihydrate, acetic acid, trehalose dihydrate and polysorbate 20 (Okrevus, concentrate for solution for infusion, Russian registration certificate number JIH[LP]-004503).

Divozilimab also relates to anti-CD20 antibodies (WHO Drug Information, Vol. 35, No. 1, 2021, CAS 2254061-60-2).

The prior art does not provide any pharmaceutical compositions comprising divozilimab as an active ingredient.

In view of the above, there is a need to develop improved stable pharmaceutical compositions of anti-CD20 antibody, in particular divozilimab, that are stable during storage in terms of "monomer content" and "acid fraction content" and that may be used as a medicinal product for the treatment of a CD20-mediated disease or disorder.

Brief description of drawings

Figure 1 is the amino acid sequence of monoclonal antibody divozilimab, showing the number of potential chemical degradation sites for light and heavy chains.

Figure 2 is monomer fraction content by size-exclusion HPLC vs. time at a temperature of +37°C.

Formulation 1 Divozilimab 25 mg/ml

Sodium acetate t/h 1.74 Acetic acid glacial to pH 5.5 D-sorbitol 50 mg/ml Polysorbate 80 0.5 mg/ml Formulation 2 Divozilimab 25 mg/ml

Sodium acetate t/h 1.74 Acetic acid glacial to pH 5.5 D-sorbitol 50 mg/ml Polysorbate 80 0.5 mg/ml L-methionine 0.15 mg/ml Formulation 3 Divozilimab 25 mg/ml

Sodium acetate t/h 1.74 Acetic acid glacial to pH 5.5 D-sorbitol 50 mg/ml Polysorbate 80 0.5 mg/ml L-methionine 1.49 mg/ml

Figure 3 is acid fraction content by ion exchange HPLC vs. time at a temperature of +37°C. Formulation 1, formulation 2 and formulation 3 have been described above in Figure 2. Figure 4 is monomer fraction content by size-exclusion HPLC vs. time at a temperature of

+25°C.

Formulation 1, formulation 2 and formulation 3 have been described above in Figure 2. Figure 5 is acid fraction content by ion exchange HPLC vs. time at a temperature of +25°C. Formulation 1, formulation 2 and formulation 3 have been described above in Figure 2. Figure 6 is monomer fraction content by size-exclusion HPLC vs. time at a temperature of +(2-8)°C.

Formulation 1, formulation 2 and formulation 3 have been described above in Figure 2. Figure 7 is acid fraction content by ion exchange HPLC vs. time at a temperature of +(2- 8)°C.

Formulation 1, formulation 2 and formulation 3 have been described above in Figure 2.

Detailed description of the invention

Definitions

Unless defined otherwise herein, all technical and scientific terms used in connection with the present invention will have the same meaning as is commonly understood by those skilled in the art.

Furthermore, unless otherwise required by context, singular terms shall include plural terms, and the plural terms shall include the singular terms.

As used in the present description and claims that follow, unless otherwise dictated by the context, the words "have", "include," and "comprise" or variations thereof such as "has", "having," "includes", "including", "comprises," or "comprising," will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers.

The term "antibody" or "immunoglobulin" (Ig) includes full-length antibodies or any antigen binding fragment (i.e., "antigen-binding portion") or individual chains thereof. The term "antibody" within the scope of the present invention is used in the broadest sense and may include, without limitation, monoclonal antibodies (including full-length monoclonal antibodies), polyclonal antibodies, humanized, fully human antibodies and chimeric antibodies.

A full-length antibody refers to a glycoprotein comprising at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds. Each heavy chain comprises a heavy chain variable region (abbreviated referred to in the present description as VH) and a heavy chain constant region. The constant region is identical in all antibodies of the same isotype, but differs in antibodies of different isotypes. Heavy chains y, a and 6 have a constant region composed of three constant domains CHI, CH2 and CH3 (in a line), and a hinge region for added flexibility (Woof J., Burton D., Nat Rev Immunol 4, 2004, pp.89-99). In mammals, known are only two types of light chains denoted by lambda (X) and kappa (K). Each light chain consists of a light chain variable region (abbreviated referred to in the present description as VL) and light chain constant region. The approximate length of a light chain is 211 to 217 amino acids. Preferably, the light chain is a lambda (X) light chain, and the constant domain CL is preferably C lambda (X).

VL and VH regions may be further subdivided into hyper-variability regions called complementarity determining regions (CDRs), located between regions that are more conserved, termed framework regions (FRs). Each VH and VL is composed of three CDRs and four FRs, arranged from amino-terminus to carboxy -terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of heavy and light chains contain a binding domain that interacts with an antigen. The constant regions of antibodies may mediate the binding of immunoglobulin to host tissues or factors, including various cells of the immune system (e.g. effector cells) and the first component (Clq) of the classical complement system.

The term "antigen-binding portion" of antibody or "antigen-binding fragment", as used in the present description, refers to one or more antibody fragments that retain the ability to specifically bind to an antigen. It has been shown that the antigen-binding function of antibody can be performed by fragments of a full-length antibody. As used in the present invention, the term "antigen-binding fragment" means a Fab-fragment, i.e. a monovalent fragment, consisting of VL, VH, CL and CHI domains, which is linked with the Fc-fragment monomer. The antibody of the present invention "which binds" a target antigen refers to an antibody that binds the antigen with sufficient affinity such that the antibody can be used as a diagnostic and/or therapeutic agent targeting a protein or cell or tissue expressing the antigen, and slightly cross-reacts with other proteins. According to analytical methods: fluorescence-activated cell sorting (FACS), radioimmunoassay (RIA) or ELISA, in such embodiments, the degree of antibody binding to a non-target protein is less than 10 % of antibody binding to a specific target protein. With regard to the binding of antibody to a target molecule, the term “specific binding” or phrases “specifically binds to” or “is specific for” a particular polypeptide or an epitope on a particular target polypeptide means binding that is significantly (measurably) different from a non-specific interaction.

The terms "anti-CD20 antibody", "antibody to CD20", "antibody specifically binding to CD20" or "antibody against CD20" and the like are interchangeable within the framework of the present invention and refer to an antibody that specifically binds to CD20.

The term "pharmaceutical composition" refers to a composition and/or formulation comprising the anti-CD20 antibody in a therapeutically effective amount and excipients or auxiliary substances (carriers, diluents, fillers, solvents, etc.), the choice and proportions of which depend on the type and route of administration and dosage.

The term "excipient" or "auxiliary substance" is used herein to describe any ingredient other than the compound(s) of the present invention. These are substances of inorganic or organic nature which are used in the pharmaceutical production/manufacturing in order to give drug products the necessary physicochemical properties.

The term "aqueous composition" as used herein refers to a water-based composition, the water in the composition may be: water, water for injections, physiologic saline (0.9%-1.0% aqueous solution of sodium chloride).

The term "lyophilized" as used herein refers to a formulation that has been subjected to a process known in the art as freeze-drying, which includes freezing the formulation followed by removal of ice from the frozen contents.

A pharmaceutical composition is "stable" if the active agent retains physical stability and/or chemical stability and/or biological activity thereof during the specified shelf life at a storage temperature, for example, of 2-8 °C. Further, the active agent may retain both physical and chemical stability, as well as biological activity. Storage period is adjusted based on the results of stability test in accelerated or natural aging conditions.

Typically, amino acids are L-amino acids. For example, if histidine and histidine hydrochloride monohydrate are used, it is typically L-histidine and L-histidine hydrochloride monohydrate. For example, if proline is used, it is typically L-proline. Amino acid equivalents, for example, pharmaceutically acceptable proline salts (for example, proline hydrochloride) may also be used.

The term "medicament" or "formulation" refers to a substance (or a mixture of substances in the form of a pharmaceutical composition) in the form of tablets, capsules, solutions, ointments and other ready forms intended for restoration, improvement or modification of physiological functions in humans and animals, and for treatment and prophylaxis of diseases, for diagnostics, anesthesia, contraception, cosmetology and others.

The term “use” applies to the possibility to use the pharmaceutical composition of anti-CD20 antibody according to the present invention to treat, relief the course of the disease or disorders, expedite the remission, reduce the recurrence rate for the diseases or disorders. The term "CD20-mediated disease or disorder" refers to any disease or disorder that is either directly, or indirectly associated with CD20, including etiology, development, progression, persistence or pathology of a disease or disorder.

"Treat", "treatment" and "therapy" refer to a method of alleviating or abrogating a biological disorder and/or at least one of attendant symptoms thereof. As used herein, to “alleviate” a disease, disorder or condition means reducing the severity and/or occurrence frequency of the symptoms of the disease, disorder, or condition. Further, references herein to "treatment" include references to curative, palliative and prophylactic treatment.

The term "parenteral administration" refers to administration regimens, typically performed by injection (infusion), and includes, in particular intravenous, intramuscular, intraarterial, intratracheal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, intraarticular, subcapsular, subarchnoid, intraspinal, epidural and intrasternal injection or infusion.

Abbreviations

VE - vertical electrophoresis

IE HPLC - ion exchange high-performance liquid chromatography.

SE HPLC - size-exclusive high-performance liquid chromatography.

PTM - post-translational modification.

ABP - acid-base profile. n/a - not applicable.

The authors of the invention have found that the presence of trehalose in the pharmaceutical compositions of divozilimab negatively affects the stability of the antibody. Further, within the framework of the present invention, it has been determined that the developed pharmaceutical composition has high stability in terms of "monomer content" and "acid fraction content".

The present invention discloses improved stable pharmaceutical compositions of anti-CD20 antibody, in particular divozilimab, that may be used as a medicinal product for the treatment of CD20-mediated diseases or disorders.

Within the framework of the present invention, it has been determined that the developed pharmaceutical composition, due to the resulting combination of excipients, has increased stability in terms of "monomer content" and "acid fraction content".

Protein aggregation is defined as the self-association of monomers in a native or partially unfolded form. Many diseases, including Alzheimer's disease, prion disorders, are associated with protein aggregation in vivo. Aggregation is also a frequent manifestation of instability observed in the production, storage and use of biologicals. Aggregation may affect efficacy and biodistribution, as well as increase chances of adverse immune reactions in patients. Adverse immune reactions to therapeutic proteins are well documented and may be clinically manifested as decreased efficacy of a medicinal product, infusion reactions, cytokine release syndrome, anaphylaxis or even death (Moussa EM, Panchai JP, Moorthy BS, Blum JS, Joubert MK, Narhi LO, Topp EM. Immunogenicity of Therapeutic Protein Aggregates. J Pharm Sci. 2016 Feb).

According to Ph. Eur. (European Pharmacopoeia), the isoform profile should be analyzed. Anionic modifications, causing a decrease in pl by 1-2 units, were shown to decrease plasma clearance and tissue accumulation. (Ryman JT, Meibohm B. Pharmacokinetics of Monoclonal Antibodies. CPT Pharmacometrics Sy st Pharmacol. 2017 Sep). These indicators are critical. While selecting the formulation, we took into account the purpose, route of administration and tolerability of the drug product (for example, reduction of discomfort during administration), as well as the stability and preservation of activity of protein molecule within the formulation.

In one aspect, the present invention relates to a pharmaceutical composition of anti-CD20 antibody comprising:

(i) anti-CD20 antibody;

(ii) acetate buffer;

(iii) sorbitol;

(iv) methionine;

(v) water for inj ection.

The concentration of anti-CD20 antibody contained in the pharmaceutical compositions of the present invention may vary depending on the desired properties of the compositions, as well as on the particular conditions, methods and purposes of use of the pharmaceutical compositions.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 1.5-120.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 1.5-1000.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 1.5-80.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 1.5-60.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 5.0-50.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 10.0-50.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 10.0-40.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 15.0-35.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 20.0-30.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 22.0-27.0 mg/ml.

In some embodiments of the invention, the anti-CD20 antibody is present at a concentration of 25.0 mg/ml.

The anti-CD20 antibody may be an antibody that specifically binds to CD20. The anti-CD20 antibody may be a full-length antibody or antigen-binding fragment thereof that specifically binds to CD20. The anti-CD20 antibody may be of different specificities (e.g. monospecific, bispecific antibody), different valencies (e.g. monovalent, bivalent, trivalent antibody), different formats (e.g. classical antibody, scFv, scFv-Fc, Minibody), different origins (e.g. murine, human, camel, chimeric antibody).

Antibodies according to the invention may be of any class (e.g. IgA, IgD, IgE, IgG, and IgM), or subclass (isotype) (e.g. IgGl, IgG2, IgG3, IgG4, IgAl and IgA2).

The anti-CD20 antibody may be, for example, rituximab, obinutuzumab, ofatumumab, ocrelizumab and others. In some embodiments of the invention, the anti-CD20 antibody is divozilimab.

In some embodiments of the invention, the acetate buffer is a mixture of sodium acetate and acetic acid.

In some embodiments of the invention, sodium acetate is present at a concentration of 0.5-

3.0 mg/ml.

In some embodiments of the invention, sodium acetate is present at a concentration of 1.0- 2.5 mg/ml.

In some embodiments of the invention, sodium acetate is present at a concentration of 1.0-

2.0 mg/ml.

In some embodiments of the invention, sodium acetate is present at a concentration of 1.5-

2.0 mg/ml.

In some embodiments of the invention, sodium acetate is present at a concentration of 1.7-

1.8 mg/ml.

In some embodiments of the invention, sodium acetate is present at a concentration of 1.74 mg/ml.

In some embodiments of the invention, sodium acetate is sodium acetate trihydrate.

In some embodiments of the invention, acetic acid is added to pH 5.0 - 6.0.

In some embodiments of the invention, acetic acid is added to pH 5.4 - 5.6.

In some embodiments of the invention, acetic acid is added to pH 5.5.

In some embodiments of the invention, acetic acid is glacial acetic acid.

In some embodiments of the invention, sorbitol is present at a concentration of 40.0-60.0 mg/ml.

In some embodiments of the invention, sorbitol is present at a concentration of 45.0-55.0 mg/ml.

In some embodiments of the invention, sorbitol is present at a concentration of 50.0 mg/ml.

In some embodiments of the invention, methionine is present at a concentration of 0.05-0.30 mg/ml.

In some embodiments of the invention, methionine is present at a concentration of 0.10-0.20 mg/ml.

In some embodiments of the invention, methionine is present at a concentration of 0.15 mg/ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody;

(ii) acetate buffer being a mixture of sodium acetate 0.5-3.0 mg/ml and acetic acid to pH 5.0-6.0;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, 1 pharmaceutical composition comprises:

(i) anti-CD20 antibody;

(ii) acetate buffer being a mixture of sodium acetate trihydrate 0.5-3.0 mg/ml and acetic acid to pH 5.0-6.0;

(iii) sorbitol 40.0-60.0 mg/ml; (iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 1.5-120.0 mg/ml;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 1.5-120.0 mg/ml;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 5.0-50.0 mg/ml;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 5.0-50.0 mg/ml;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 20.0-30.0 mg/ml;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 20.0-30.0 mg/ml;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 25.0 mg/ml;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 25.0 mg/ml;

(iii) sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody;

(ii) acetate buffer being a mixture of sodium acetate 1.0-2.0 mg/ml and acetic acid to pH 5.4-5.6;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 1.5-120.0 mg/ml;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, 1 pharmaceutical composition comprises:

(i) anti-CD20 antibody 5.0-50.0 mg/ml;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml. In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 20.0-30.0 mg/ml;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 25.0 mg/ml;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(ii) anti-CD20 antibody;

(iii) acetate buffer being a mixture of sodium acetate 1.5-2.0 mg/ml and acetic acid to pH 5.4-5.6;

(vi) sorbitol 45.0-55.0 mg/ml;

(vii) methionine 0.10-0.20 mg/ml;

(viii) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody;

(ii) acetate buffer being a mixture of sodium acetate trihydrate 1.5-2.0 mg/ml and acetic acid to pH 5.4-5.6;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(ii) anti-CD20 antibody 1.5-120.0 mg/ml;

(vi) sorbitol 45.0-55.0 mg/ml;

(vii) methionine 0.10-0.20 mg/ml;

(viii) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 1.5-120.0 mg/ml;

(ii) acetate buffer being a mixture of sodium acetate trihydrate 1.5-2.0 mg/ml and acetic acid to pH 5.4-5.6; (iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 5.0-50.0 mg/ml;

(ii) acetate buffer being a mixture of sodium acetate 1.5-2.0 mg/ml and acetic acid to pH 5.4-5.6;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 5.0-50.0 mg/ml;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 20.0-30.0 mg/ml;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 20.0-30.0 mg/ml;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 25.0 mg/ml;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 25.0 mg/ml; (ii) acetate buffer being a mixture of sodium acetate trihydrate 1.5-2.0 mg/ml and acetic acid to pH 5.4-5.6;

(iii) sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody;

(ii) acetate buffer being a mixture of sodium acetate 1.74 mg/ml and acetic acid to pH 5.5;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody;

(ii) acetate buffer being a mixture of sodium acetate trihydrate 1.74 mg/ml and acetic acid to pH 5.5;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 1.5-120.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 1.5-120.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, 1 pharmaceutical composition comprises:

(i) anti-CD20 antibody 1.5-60.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml; (v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 1.5-60.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 5.0-50.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 5.0-50.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 10.0-40.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, 1 pharmaceutical composition comprises:

(i) anti-CD20 antibody 10.0-40.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, 1 pharmaceutical composition comprises:

(i) anti-CD20 antibody 20.0-30.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, 1 pharmaceutical composition comprises:

(i) anti-CD20 antibody 20.0-30.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, 1 pharmaceutical composition comprises:

(i) anti-CD20 antibody 25.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) divozilimab 25.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) anti-CD20 antibody 25.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml.

In some embodiments of the invention, the pharmaceutical composition comprises:

(i) divozilimab 25.0 mg/ml;

(iii) sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for inj ection to 1 ml. In one aspect, the present invention relates to a pharmaceutical composition of anti-CD20 antibody, which is provided in dry (i.e. powder or granular) form for reconstitution in a suitable solvent (e.g. water) prior to administration. Such formulation may be prepared by, for example, lyophilisation, i.e. a process which is known in the art as freeze drying, and which involves freezing a product followed by removal of solvent from frozen material.

In one aspect, the present invention relates to a pharmaceutical composition of anti-CD20 antibody produced by lyophilization of any of the above pharmaceutical compositions of anti-CD20 antibody. Accordingly, the pharmaceutical compositions according to the present invention may be either aqueous pharmaceutical compositions or lyophilized pharmaceutical compositions (lyophilizates).

Lyophilizates are used to produce other dosage forms. For example, a lyophilizate for producing an injectable solution, a lyophilizate for producing a concentrate for producing an injectable solution. Lyophilizates are reconstituted by dissolving same in a suitable solvent, most typically in water for injection. Also, lyophilized compositions are first reconstituted in the required volume of solvent (most typically in water) and then further diluted in a suitable solvent (e.g. 5% glucose solution, 0.9% sodium chloride solution).

The pharmaceutical compositions according to the present invention are typically suitable for parenteral administration as sterile formulations intended for administration in a human body through the breach in skin or mucosal barriers, bypassing the gastrointestinal tract by virtue of injection, infusion and implantation. In particular, it is contemplated that parenteral administration includes, inter alia, subcutaneous, intraperitoneal, intramuscular, intravenous, intraarterial, intrathecal, intraventricular, intraurethral, intracranial, intrasynovial, transdermal injection or infusion; and kidney dialytic infusion techniques. Preferred embodiments include intravenous and subcutaneous routes. Any method for administering peptides or proteins accepted in the art may be suitably employed for the composition of anti-CD20 antibody according to the present invention.

The pharmaceutical composition of anti-CD20 antibody according to the present invention may be used after dilution. To this end, the required volume of the composition is transferred from a vial to an infusion container comprising a sterile 0.9% sodium chloride solution or a sterile 5% dextrose solution. The resulting solution is stirred by gently turning the infusion container over.

The pharmaceutical compositions according to the present invention may be stored in any suitable container. For example, a glass or plastic container, vial, ampoule, syringe, cartridge, or bottle of the desired volume. The containers may be provided with additional means for administration, for example, droppers, auto-injectors.

A pharmaceutical composition according to the invention may be manufactured, packaged, or widely sold in the form of a single unit dose or a plurality of single unit doses in the form of a ready formulation. The term "single unit dose" as used herein refers to discrete quantity of a pharmaceutical composition containing a predetermined quantity of an active ingredient. The quantity of an active ingredient typically equals the dose of the active ingredient to be administered in a subject, or a convenient portion of such dose, for example, half or a third of such dose.

In one aspect, the present invention relates to the use of the above pharmaceutical composition of anti-CD20 antibody for the treatment of a CD20-mediated disease or disorder in a subject in need thereof.

In some embodiments of the invention, the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 125 mg.

In some embodiments of the invention, the pharmaceutical composition is administered at a dose of anti-C.D20 antibodv of 500 mu In some embodiments of the invention, the pharmaceutical composition is administered every 24 weeks.

In some embodiments of the invention, the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 125 mg every 24 weeks.

In some embodiments of the invention, the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 500 mg every 24 weeks.

In some embodiments of the invention, the use of the above pharmaceutical composition of anti-CD20 antibody for the treatment of a CD20-mediated disease or disorder in a subject in need thereof comprises at least 5 cycles of administration of the pharmaceutical composition.

In some embodiments of the invention, the first dose of anti-CD20 antibody of 125 mg or 500 mg may be administered to the subject in two steps with a 14-day interval at a dose of 62.5 mg or 250 mg (i.e., on day 1, the anti-CD20 antibody is administered to a subject at a dose of 62.5 mg or 250 mg; on day 15, the anti-CD20 antibody is administered again at analogous doses of 62.5 mg or 250.0 mg). The anti-CD20 antibody is administered in the form of the above pharmaceutical composition.

In some embodiments of the invention, the use of the above pharmaceutical composition of anti-CD20 antibody for treating a CD20-mediated disease or disorder in a subject in need thereof includes at least 5 cycles of administration of the pharmaceutical composition, where cycle 1 includes the first administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 62.5 mg and the second administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 62.5 mg 2 weeks following the first administration; the following cycles include the administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 125 mg every 24 weeks following the first administration.

In some embodiments of the invention, the use of the above pharmaceutical composition of anti-CD20 antibody for treating a CD20-mediated disease or disorder in a subject in need thereof includes at least 5 cycles of administration of the pharmaceutical composition, where cycle 1 includes the first administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 250 mg and the second administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 250 mg 2 weeks following the first administration; the following cycles include the administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 500 mg every 24 weeks following the first administration.

In some embodiments of the invention, the anti-CD20 antibody is divozilimab.

In some embodiments of the invention, the above pharmaceutical composition is administered at a dose of divozilimab of 125 mg.

In some embodiments of the invention, the above pharmaceutical composition is administered at a dose of divozilimab of 500 mg.

In some embodiments of the invention, the above pharmaceutical composition of divozilimab is administered every 24 weeks.

In some embodiments of the invention, the above pharmaceutical composition is administered at a dose of divozilimab of 125 mg every 24 weeks.

In some embodiments of the invention, the above pharmaceutical composition is administered at a dose of divozilimab of 500 mg every 24 weeks.

In some embodiments of the invention, the use of the above pharmaceutical composition of divozilimab for the treatment of a CD20-mediated disease or disorder in a subject in need thereof comprises at least 5 cycles of administration of the above pharmaceutical composition of divozilimab.

In some embodiments of the invention, the use of the above pharmaceutical composition of divozilimab for the treatment of a CD20-mediated disease or disorder in a subject in need thereof includes at least 5 cycles, where cycle 1 includes the first administration of the above pharmaceutical composition at a dose of divozilimab of 62.5 mg and the second administration of the above pharmaceutical composition at a dose of divozilimab of 62.5 mg 2 weeks following the first administration; the following cycles include the administration of the above pharmaceutical composition at a dose of divozilimab of 125 mg every 24 weeks following the first administration.

In some embodiments of the invention, the use of the above pharmaceutical composition of divozilimab for the treatment of a CD20-mediated disease or disorder in a subject in need thereof includes at least 5 cycles, where cycle 1 includes the first administration of the above pharmaceutical composition at a dose of divozilimab of 250 mg and the second administration of the above pharmaceutical composition at a dose of divozilimab of 250 mg 2 weeks following the first administration; the following cycles include the administration of the above pharmaceutical composition at a dose of divozilimab of 500 mg every 24 weeks following the first administration.

In some embodiments of the invention, the CD20-mediated disease or disorder is selected from the group:

(i) oncological disease or disorder or

(ii) autoimmune disease or disorder.

In some embodiments of the invention, the oncological disease or disorder is selected from the group: B-cell lymphoma, leukemia.

In some embodiments of the invention, B-cell lymphoma is non-Hodgkin's lymphoma, Hodgkin's lymphoma.

In some embodiments of the invention, leukemia is selected from the group: chronic lymphocytic leukemia, small lymphocytic lymphoma.

In some embodiments of the invention, the autoimmune disease or disorder is selected from the group: multiple sclerosis, axonal neuropathy, ANCA-associated vasculitides, IgG4-related disease, amyloidosis, axial spondyloarthritis, antiphospholipid syndrome, Takayasu's arteritis, autoimmune aplastic anemia, autoimmune inner ear disease, autoimmune hemolytic anemia, autoimmune hyperlipidemia, autoimmune dysautonomia, autoimmune urticaria, autoimmune neuromyotonia (Isaacs syndrome), autoimmune retinopathy, autoimmune thrombocytopenic purpura, warm antibody hemolytic anemia, autoimmune testicular disease, autoimmune angioedema, autoimmune hepatitis, autoimmune diabetes, autoimmune immunodeficiency, autoimmune pericarditis, autoimmune thyroiditis, autoimmune progesterone dermatitis, Addison's disease, Behcet's disease, Graves' disease (diffuse toxic goiter), neuromyelitis optica spectrum diseases (Devic's disease), Castleman's disease, Crohn's disease, Ormond's disease (retroperitoneal fibrosis), Raynaud's disease, Churg-Strauss syndrome, bullous pemphigoid, epidermolysis bullosa, vasculitis, renal vasculitis, antineutrophil cytoplasmic antibody-associated vasculitides, vesiculobullous dermatosis, giant cell arteritis, vitiligo, lupus nephritis, inflammatory aortic aneurysm, inflammatory bowel disease, congenital heart block, pemphigus vulgaris, dermatitis herpetiformis, gestational pemphigoid, hypogammaglobulinemia, glomerulonephritis, granulomatosis with polyangiitis (Wegener's syndrome), dermatomyositis, dilated cardiomyopathy, demyelinating neuropathy, discoid lupus erythematosus, idiopathic recurrent macrohematuria (IgA nephropathy), idiopathic inflammatory myopathy, idiopathic inflammatory pseudotumor, idiopathic thrombocytopenic purpura, idiopathic hypocomplementemic tubulointerstitial nephritis, idiopathic pulmonary fibrosis, diabetes mellitus, pure red cell aplasia, Balo's concentric sclerosis, lichen planus, lichen sclerosus, leukocytoclastic vasculitis, limited scleroderma (CREST syndrome), limbic encephalitis, linear IgA bullous dermatosis, pemphigus foliaceus, Waldenstrom macroglobulinemia, mediastinal fibrosis, myasthenic crisis, Lambert-Eaton myasthenic syndrome, myasthenia gravis, microscopic polyangiitis, myositis, Coxsackie myocarditis, multiple sclerosis, multifocal motor neuropathy, multifocal fibrosclerosis, optic neuritis, undifferentiated connective tissue disease uveitis, neutropenia, ulcerative colitis, anti-tubular/glomerular basement membrane antibody-associated nephritis, opsoclonus, acute motor axonal neuropathy, acute hemorrhagic leukoencephalitis, acute disseminated encephalomyelitis, palindromic rheumatism, paraneoplastic cerebellar degeneration, paraneoplastic syndrome, paraproteinemic polyneuropathy, paroxysmal nocturnal hemoglobinuria, ocular pemphigus, mucous membrane pemphigoid, primary biliary cirrhosis, primary sclerosing cholangitis, periaortitis, periarteritis, perivenous encephalomyelitis, peripheral neuropathy, peripheral uveitis, pernicious anemia, subacute bacterial endocarditis, polymyositis, polyneuropathy with organomegaly (POEMS syndrome), transverse myelitis, Dressier's syndrome, post-traumatic pericarditis, psoriasis, psoriatic arthritis, pemphigus, Henoch- Schonlein purpura, anti -HL A antibody-mediated transplant rejection, reactive arthritis, rheumatic fever, polymyalgia rheumatica, rheumatoid arthritis, reflex sympathetic dystrophy, refractory epilepsy, relapsing polychondritis, sarcoidosis, Guillain-Barre syndrome, Kawasaki syndrome, Cogan syndrome, Mikulicz syndrome, Stiff-person syndrome, Sjogren's syndrome, Evans syndrome, systemic lupus erythematosus, systemic scleroderma, limited scleroderma, Riedel's thyroiditis, Hashimoto's thyroiditis, thyroid-associated ophthalmopathy, thyroid crisis, thrombotic thrombocytopenic purpura, polyarteritis nodosa, fetal and neonatal alloimmune thrombocytopenia, fibrosing alveolitis, B-cell non-Hodgkin's lymphoma, B-cell chronic lymphocytic leukemia, cold haemagglutinin disease, Sydenham's chorea (chorea minor or rheumatic chorea), chronic Lyme disease, chronic inflammatory demyelinating polyneuropathy, chronic recurrent multifocal osteomyelitis, chronic sclerosing sialadenitis (Kiittner's tumor), celiac disease, endometriosis, encephalitis, autoantibody-associated encephalopathy, Hashimoto's encephalitis, eosinophilic fasciitis, essential mixed cryoglobulinemia, juvenile diabetes, juvenile rheumatoid arthritis.

In some embodiments of the invention, multiple sclerosis is remitting multiple sclerosis, secondary progressive multiple sclerosis, primary progressive multiple sclerosis, multiple sclerosis with exacerbations, highly active multiple sclerosis, aggressive multiple sclerosis, malignant multiple sclerosis (Marburg's variant), myelinoclastic diffuse Schilder's sclerosis, myelocortical multiple sclerosis, a clinically isolated syndrome.

In some embodiments of the invention, the subject of treatment, or patient, is a mammal, preferably a human subject. Said subject may be either male or female, of any age.

In one aspect, the present invention relates to a method of treating a CD20-mediated disease or disorder comprising administering the above pharmaceutical composition to a subject in need thereof in a therapeutically effective amount.

In some embodiments of the invention, the above pharmaceutical composition is administered at a dose of anti-CD20 antibody of 125 mg.

In some embodiments of the invention, the above pharmaceutical composition is administered at a dose of anti-CD20 antibody of 500 mg.

In some embodiments of the invention, the above pharmaceutical composition is administered every 24 weeks.

In some embodiments of the invention, the above pharmaceutical composition is administered at a dose of anti-CD20 antibody of 125 mg every 24 weeks.

In some embodiments of the invention, the above pharmaceutical composition is administered at a dose of anti-CD20 antibody of 500 mg every 24 weeks.

In some embodiments of the invention, the method of treating a CD20-mediated disease or disorder includes at least 5 cycles of administration of the above pharmaceutical composition. In some embodiments of the invention, the first dose of anti-CD20 antibody of 125 mg or 500 mg may be administered to the subject in two steps with a 14-day interval at a dose of 62.5 mg or 250 mg (i.e., on day 1, the anti-CD20 antibody is administered to a subject at a dose of 62.5 mg or 250 mg; on day 15, the anti-CD20 antibody is administered again at analogous doses of 62.5 mg or 250.0 mg). The anti-CD20 antibody is administered in the form of the above pharmaceutical composition.

In some embodiments of the invention, the method of treating a CD20-mediated disease or disorder includes at least 5 cycles, where cycle 1 includes the first administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 62.5 mg and the second administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 62.5 mg 2 weeks following the first administration; the following cycles include the administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 125 mg every 24 weeks following the first administration.

In some embodiments of the invention, the method of treating a CD20-mediated disease or disorder includes at least 5 cycles, where cycle 1 includes the first administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 250 mg and the second administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 250 mg 2 weeks following the first administration; the following cycles include the administration of the above pharmaceutical composition at a dose of anti-CD20 antibody of 500 mg every 24 weeks following the first administration.

In some embodiments of the invention, the anti-CD20 antibody is divozilimab.

In some embodiments of the invention, the method of treating a CD20-mediated disease or disorder includes at least 5 cycles of administration of the above pharmaceutical composition of divozilimab.

In some embodiments of the invention, the method of treating a CD20-mediated disease or disorder includes at least 5 cycles, where cycle 1 includes the first administration of the above pharmaceutical composition at a dose of divozilimab of 62.5 mg and the second administration of the above pharmaceutical composition at a dose of divozilimab of 62.5 mg 2 weeks following the first administration; the following cycles include the administration of the above pharmaceutical composition at a dose of divozilimab of 125 mg every 24 weeks following the first administration.

In some embodiments of the invention, the method of treating a CD20-mediated disease or disorder includes at least 5 cycles, where cycle 1 includes the first administration of the above pharmaceutical composition at a dose of divozilimab of 250 mg and the second administration of the above pharmaceutical composition at a dose of divozilimab of 250 mg 2 weeks following the first administration; the following cycles include the administration of the above pharmaceutical composition at a dose of divozilimab of 500 mg every 24 weeks following the first administration.

(i) oncological disease or disorder or

(ii) autoimmune disease or disorder.

The pharmaceutical compositions may be administered as a single therapeutic agent or in combination with additional therapeutic agents as needed. Thus, in one embodiment, the present methods for treatment and/or prophylaxis are used in combination with administration of a therapeutically effective amount of another active agent. The other active agent may be administered before, during or following the administration of the pharmaceutical compositions according to the present invention. The other active agent may be administered as part of the present composition or, alternatively, as a separate formulation.

Implementation of the invention

The following examples are provided for better understanding of the invention. These examples are for purposes of illustration only and are not to be construed as limiting the scope of the invention in any manner.

All publications, patents, and patent applications cited in this specification are incorporated herein by reference. Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be readily apparent to those of ordinary skill in the art in light of the teachings of this invention that certain changes and modifications may be made thereto without departing from the spirit or scope of the appended embodiments.

Methods.

1. Protein concentration assay in test samples.

Protein concentration was determined by UV spectrophotometry at a wavelength of 280 nm in UV spectrophotometry plates.

Each sample was diluted with the appropriate solution of excipients to a concentration of ~ 0.5 mg/ml. 150 pl of the diluted sample was placed into a well of UV spectrophotometry plate. Optical density of solutions in the plate wells was measured using a plate spectrophotometer at a wavelength of 280 nm. A corresponding solution of excipients was used as a reference solution.

Concentration (mg/ml) of protein (C) was calculated using the following formula:

C= A(280)*bs*lC= A280*bc*l,

A280 is the value of optical density at a wavelength of 280 nm; e is the extinction coefficient of test protein; b is the total dilution factor for a sample;

1 is layer thickness in a plate well (for 150 pl, 1 = 0.42 cm; for a half-area plate, 175 pl = 1 cm).

2. Buffer solution replacement and sample concentration.

Samples were dialyzed and concentrated in multiple manners:

- in Stirred Cell (Millipore) concentration cells under pressure,

- in Amicon centrifuge tubes with a 10 kDa membrane (Millipore),

- in dialysis tubes with a 2 ml or 0.5 ml 10 kDa membrane (Thermo scientific).

Dialysis was performed for 2-3 hours, thereafter the used dialysis buffer was replaced with a new one, dialysis was resumed for additional several hours while constantly stirring in an orbital shaker (100 rpm) and at a room temperature.

3. Evaluation of colloidal stability by shake test.

Test samples were divided into 2 portions of 200 pl each and placed into glass vials, 1 vial per formulation was transferred to a refrigerator for aging at 2-8 °C, the remaining vials were placed in a shaker and shaken at 800 rpm at 2-8 °C for the specified period. After the stress, the vials were removed from the shaker and transferred for analysis.

4. Evaluation of colloidal stability by cryoconcentration.

Test samples were divided into 2 portions and placed into plastic vials: 1 vial per formulation was stored in a refrigerator at 2-8 °C; the remaining vials were stored in a freezer at minus 16-20 °C for the specified period of time. After the stress, the vials were removed from the freezer, kept at room temperature until the contents were completely thawed; the solutions were mixed and transferred for analysis.

5. Evaluation of thermal stability by "Thermostress"

Test samples were divided into 2 portions and placed into separate glass vials: 1 vial per composition was stored in a refrigerator at 2-8 °C, the remaining vials were incubated in a thermostat at a required temperature for the specified period of time. After heating, the vials were removed from the thermostat, kept at room temperature for about 15 minutes, and transferred for analysis.

6. Determination of protein homogeneity and aggregation point by dynamic light scattering (DLS).

The homogeneity of test samples was determined using the DynaPro Plate Reader II (Wyatt). To this end, 35 ml of the solution was added to a well of a 384-well 384 LVZEB plate. After filling, the plate was sealed with a film. To remove air bubbles from the wells after filling, the plate on a lint-free wipe was centrifuged in a plate rotor for 3-5 minutes, 3000 rpm; during this, the temperature in the centrifuge chamber should not be below 20 °C to avoid sample condensation on the film.

Measurement temperature: 25 °C.

Keeping for 5 seconds at the temperature before starting the measurement.

Scattered light intensity at 0 = 158°. Number of measurements per replicate - 10.

Time per measurement - 5 s.

The aggregation point of test proteins was determined using the DynaPro Plate Reader II (Wyatt). The samples were prepared according to the above method for determining homogeneity by DLS.

Heating temperature range - 40-85 °C.

Heating rate - 0.188 °C/min.

Number of samples per measurement - no more than 16.

Scattered light intensity - 0 = 158°.

Number of measurements per replicate - 3.

Time per measurement - 5 s.

We built a temperature trend using the device software, which also automatically calculates a protein aggregation point (Onset temperature).

7. Determination of samples purity by size-exclusion high-performance liquid chromatography (SEC HPLC).

Tosoh column TSK-GelG3000SWXL 7.8 mm ID x 30 cm, cat. No 08541.

Column temperature: 25 °C

Mobile phase flow rate: 0.7 ml/min

Injection volume: 10 pl (sample concentration - 5 mg /ml. dilution - with an excipient solution). For other concentrations, the injection volume is equivalent to the concentration.

Sample concentration: 5 mg/ml.

Detector wavelength: 220 nm.

Elution time: 25 min.

Mobile phase: Disodium hydrogen phosphate anhydrous 7.1 mg/ml.

Sodium chloride 17.54 mg/ml.

The mobile phase pH was adjusted to 7.0 with orthophosphoric acid.

8. Determination of acid-base profile of samples on Caliper LabChip GX II.

Preparation of samples.

Samples were diluted to a concentration of 1 mg/ml. 2 pl of carboxypeptidase B (CpB) solution at a concentration of 5 mg/ml was added to 200 pl of the resulting solution. The solution was mixed and incubated for 2 hours at 37°C. Test samples were dialyzed against three changes of water in Amicon Ultra centrifuge tubes and concentrated to 2 mg/ml.

Preparation of working solutions.

Working solutions and a plate with test samples were prepared in accordance with the manufacturer's protocol using the HT Protein Charge Variant Labeling Kit.

Preparation of chip.

The chip and the buffer tube were prepared according to the manufacturer's protocol using buffer solutions from the Protein Charge Variant Buffer Kit.

Launch of the analysis is a standard procedure. Protein Charge Variant 68s was used as a method of analysis.

9. Determination of acid-base profile of samples by ion-exchange (IE) HPLC.

Chromatographic conditions:

Column: Pro PACTM WCX-10 Analytical (Dionex), 250 x 4 mm;

Precolumn: Pro Pac WCX-10G, 4 x 50 mm;

Mobile phase: Solution A 0.02 M 2-(N-morpholino)ethanesulfonic acid (MES), pH 6.0;

Solution B 0.02 M MES, 0.4 M sodium chloride, pH 6.0;

Mobile phase speed: 0.7 ml/min;

Column temperature: 30°C;

Autosampler temperature: 5°C;

Detector: UV, 280 nm;

Injected sample volume: 40 pl.

10. PTM in silico.

The protein structure may contain certain regions prone to chemical degradation, for example, acetylation, carb amyl ati on, methylation, phosphorylation, oxidation and other processes. The types of chemical modifications are determined by the structure of amino acids or their sequence in the polypeptide chain. To predict the proneness of proteins towards the above modifications, PTM sites and their number may be studied in silico in the primary structure of protein.

11. Stability study.

The test samples were divided into several aliquots and placed into distinct sterile glass vials: 1 vial for each control point was placed in a thermostat, incubated at 25 °C / 37 °C for 6 months / 4 weeks and at (5 ± 3) °C for 24 months, with periodic collection of control points according to the schedule. While being collected at control points and following storage, the vials were removed from a thermostat or refrigerator and transferred for analysis.

12. Determination of purity by reducing and non-reducing vertical gel electrophoresis in polyacrylamide gel (PAG)

The determination was carried out by vertical electrophoresis in polyacrylamide gel (PAAG) under reducing and non-reducing conditions (10 and 40 pg loaded) in the presence of sodium lauryl sulfate, followed by gel staining with acid blue dye 83.

The determination was carried out using an electrophoresis apparatus under the following parameters: gel width of at least 13 cm, height (from the lower edge of the comb tooth) of at least 12 cm, thickness of 1 mm. To form the wells in the stacking gel, we used a comb with at least 20 teeth, dimensions: width of 4-6.5 mm, thickness of 1 mm.

The substance solution was diluted with water to a concentration of 2 mg/ml.

80 pl of the resulting solution was mixed with 80 ml of a sample application buffer solution containing 2-mercaptoethanol (reducing conditions) or not containing 2-mercaptoethanol (nonreducing conditions). The resulting mixtures were incubated at 100 °C for 1 min (samples not containing 2-mercaptoethanol) or 3 min (samples containing 2-mercaptoethanol), cooled down to room temperature, then centrifuged for 4-6 s, and mixed (test solutions).

Reducing electrophoresis was performed in 12.5% PAAG, non-reducing electrophoresis was performed in 8% PAAG.

The lower reservoir of the electrophoresis apparatus was filled with an electrode buffer solution, an electrophoretic cell with gel was inserted into the chamber. The upper reservoir was filled with an electrode buffer solution, air bubbles were removed from the wells.

Reducing and non-reducing electrophoresis was performed at room temperature in the constant voltage mode. The voltage is 100 V while the bromophenol blue dye front migrates through the stacking gel. After the bromophenol blue dye front entered the lower resolving gel by 5-7 mm, the voltage was increased to 180 V. After the blue bromophenol dye reached 10 cm from the lower edge of the stacking gel, the voltage was turned off, the gel was separated from the cell glass. After non-reducing electrophoresis, the gel was incubated with a fixing solution for 16-18 hours. The solution was then drained, the staining solution was added, and the mixture was incubated on an orbital shaker at 50 rpm for 1 h. The staining solution was then drained, the washing solution was heated to a temperature of 40-50 °C, the gel was poured, and the mixture was placed on an orbital shaker at 50 rpm for 30 min. The washing procedure was repeated until the bands in the sensitivity solution track were clearly visualized. The stained gel was scanned.

The number of main diffuse bands observed in the gel after reducing and non-reducing electrophoresis was determined by the tracks of solutions containing 10 pg of protein.

The substance purity was assessed by the tracks of solutions containing 40 pg of protein in a sample application buffer solution containing or not containing 2-mercaptoethanol using the GelPro software.

Examples

The examples described below are given for the anti-CD20 antibody divozilimab.

Example 1.

The critical parameters of the acid-base profile and of aggregate content are influenced by the excipient formulation. This example studies the influence of stabilizers and osmotic agents on test quality indicators.

For the study, we used osmotic agents at concentrations providing osmotic pressure in the blood flow: 270-330 mOsm. The following excipients were subjected to screening: sodium acetate trihydrate (buffering agent, maintaining a required pH level), glacial acetic acid (maintaining a required pH level), D-trehalose dihydrate (stabilizer, osmolyte), D-sorbitol (stabilizer, osmolyte), D-mannitol (stabilizer, osmolyte), L-proline (stabilizer, osmolyte).

Excipient formulations for test pharmaceutical compositions are shown in Table 1.

Table 1 - Test pharmaceutical compositions

The study was carried out for samples containing 5 mg/ml of divozilimab. Samples with various excipient formulations were prepared by diafiltration in accordance with method 2. In order to identify the effect of various excipients, samples in test formulations were subjected to stresses in accordance with methods 3, 4, 5. Before and following stress, the samples were analyzed in accordance with methods 6, 7, 8. The results are shown in Tables 2, 3 and 4. Table 2 - SE HPLC results summary for samples before and after stresses

Table 3 - Homogeneity results summary by dynamic light scattering before and after stresses

Table 4 - Results summary for acid-base profile of samples before and after stresses

** The absolute change was calculated by the formula:

A =|acid fraction content before stress-acid fraction content following stress|+|basic fraction content before stress-basic fraction content following stress|+ (dominating fraction content before stress-dominating fraction content following stress] According to the results of SE HPLC, the samples supplemented with sorbitol have the smallest increase in aggregates after thermostress and shake test. The formulation containing trehalose dihydrate as an osmotic agent shows the greatest change in the acid-base profile and decreased monomer content after thermostress; it is not recommended for further development.

According to the results of dynamic light scattering and according to the value of the aggregation point, all samples show high thermal stability. It can also be observed that adding an osmotic agent increases the thermal stability of the protein in the solution (the first sample free of an osmotic agent showed the lowest aggregation point value in the group). When comparing the hydrodynamic radius of compositions containing sorbitol, there is a tendency to greater compaction of the protein globule.

The optimal stabilizer was selected using the method for in silico prediction of PTMs described in method 10. According to the results of the primary amino acid sequence analysis, protein degradation may potentially take place via the pathway of deaimidation of the side groups of some amino acids (for example, asparagine and glutamine) and methionine oxidation processes. Figure I shows the number of potential chemical degradation sites for light and heavy chains.

The light chain contains potentially two isomerization sites and one deamidation site. The heavy chain contains one hydrolysis and glycosylation site, three isomerization sites, six deamidation sites, and seven oxidation sites.

According to the results of the analysis of the primary protein sequence of divozilimab, protein degradation may potentially take place via the pathway of deaimidation of the side groups of some amino acids (for example, asparagine and glutamine) and methionine oxidation processes.

L-methionine was proposed to be used as a stabilizer preventing oxidation processes. This excipient added to the pharmaceutical composition acts as a stabilizer, an absorber of dissolved oxygen and an absorber of free radicals. It competes with oxidation-sensitive regions within a molecule for oxidizing agents.

To select a stabilizer within the pharmaceutical composition, L-methionine in various concentrations and polysorbate 80 were subjected to screening.

Excipient formulations for test pharmaceutical compositions are shown in Table 5.

Table 5 - Test pharmaceutical compositions

The study was carried out for samples containing 25 mg/ml of divozilimab. Samples with various excipient formulations were prepared by diafiltration in accordance with method 2. To study 11 at temperatures of 37°C, 25°Cand (2-8)°C for 5 weeks, 6 months and 24 months, respectively. Before and following stress, the samples were analyzed in accordance with methods 7 and 9. The results are shown in Tables 6, 7 and 8.

Table 6 - Stability results summary for samples stored at +37°C for 5 weeks

Table 7 - Stability results summary for samples stored at +25°C for 6 months

Table 8 - Stability results summary for samples stored at +(2-8) for 24 months°C

The monomer content by SE HPLC and acid fractions by IE HPLC vs. all temperature regimes are shown in Figures 2, 3, 4, 5, 6 and 7. The product of formulation 2 has a smaller decrease in the monomer content by SE HPLC and a smaller increase in acid fractions under all temperature regimes during storage.

Example 2.

The selected formulation of excipients was subjected to storage according to method 11 for temperature regimes of 25 °C, 37°Cand (2-8) °C for 5 weeks, 6 months and 24 months, respectively, at divozilimab concentrations of 25 mg/ml and 50 mg/ml.

Stability results were obtained according to methods 1, 7, 9, 12 and are shown in Tables 9, 10 and 11 for divozilimab concentrations of 25 mg/ml and 50 mg/ml.

Table 9 - Stability results summary for samples stored at +37°C for 5 weeks.

Table 10 - Stability results summary for samples stored at +25°C for 6 months.

Table 11 - Stability results summary for samples stored at +(2-8)°C for 24 months.

* The change was calculated by the formula:

A = (fraction content at the control point - fraction content at the input control)

** The absolute change was calculated by the formula:

A abs=|acid fraction content before stress-acid fraction content following stress|+|basic fraction content before stress-basic fraction content following stress|+ (dominating fraction content before stress-dominating fraction content following stress]

The stability data for samples with divozilimab concentrations of 25 mg/ml and 50 mg/ml are comparable. Example 3. Use of divozilimab for treating patients with multiple sclerosis with exacerbations, phase II trial, efficacy and safety study of BCD-132-2.

Trial design

Study no. BCD-132-2 in its design was an international, multicenter, randomized, doubleblind, double-masked, placebo-controlled trial using an active comparator (teriflunomide).

The trial No. BCD-132-2 assessed the efficacy of divozilimab (BCD-132) in treating patients with multiple sclerosis with exacerbations by brain MRI indicators, as well as by clinical indicators associated with exacerbations and the level of disability following administration of divozilimab at a dose of 125 and 500 mg compared with teriflunomide and compared with placebo. The study included patients diagnosed with multiple sclerosis (according to 2017 McDonald criteria for the diagnosis of multiple sclerosis), type of course - multiple sclerosis with exacerbations, and the level of disability by the EDSS scale of not higher than 5.5.

Following stratification, patients were randomized to the following arms of the trial: 125 mg divozilimab arm, 500 mg divozilimab arm, teriflunomide arm and placebo arm in a ratio of 2:2:2: 1, respectively. The trial enrolled a total of 319 subjects at the screening stage, whereas 271 participants were randomized to the trial. In the main study period, 24 weeks of therapy (1 cycle of therapy), patients of the trial product arms received divozilimab in corresponding dosages (one arm at a dose of 125 mg and the other arm at a dose of 500 mg) via intravenous infusion. Further, the patients received the first pre-specified dose within the first cycle (125 or 500 mg) via two intravenous infusions with a 14-day interval (i.e., on Day 1, patients received intravenous infusion of divozilimab at a dose of 62.5 mg or 250 mg depending on the allocation arm; on Day 15, the same doses were infused again). Also, all patients randomized to one of the two trial product arms, in order to maintain the double-blind design, received an oral placebo in tablet form once a day. Patients of the comparison arm received oral teriflunomide daily at a dose of 14 mg 1 time a day. Further, in order to maintain the double-blind design, patients of the comparator arm received intravenous placebo infusions at a frequency and dosage regimen analogous to those of the trial divozilimab arms. Patients of the placebo arm received intravenous placebo infusions at a frequency and dosage regimen analogous to those of the trial divozilimab arms (i.e., infusions on Days 1 and 15); they also received oral placebo in tablet form once a day.

The data of the 1st milestone of the trial (24 weeks of therapy) were obtained and analyzed. Clinical efficacy results summary from trial No. BCD-BCD- 132-2.

The results demonstrated superiority of the trial divozilimab at both doses (125 mg and 500 mg) over placebo and its noninferiority to the comparator teriflunomide in the primary endpoint that reflects the activity of the demyelinating process in the brain in patients with multiple sclerosis with exacerbations and that is the most reliable value for assessing the therapy response during the analyzed period (6 months) according to the EMA guidelines on clinical investigation of medicinal products for the treatment of multiple sclerosis - "Total T1 Gd+ lesions (per scan) detected by brain MRI following 24 weeks of blinded treatment with divozilimab/teriflunomide/placebo".

The trial was based on two hypotheses: the hypothesis of the superiority of the trial product over placebo and the hypothesis of non-inferior efficacy between the trial product BCD- 132 and the comparator teriflunomide. These hypotheses were verified for the efficacy endpoint, "Total T1 Gd+ lesions (per scan) detected by brain MRI following 24 weeks of blinded treatment with BCD- 132/teriflunomide/placebo". The one-sided 97.5% confidence interval for the difference in arithmetic means of total T1 Gd+ lesions per scan between the BCD-132, 125 mg group and the placebo group was (-Inf ; -0.6075], that between the BCD-132, 500 mg group and placebo group was (-Inf ; -0.5974], The results in the groups of the trial BCD-132 indicate that the upper limit of the confidence interval in both groups does not exceed the preset limit of superiority 0. The two- sided 95% confidence interval for the difference in arithmetic means of total T1 Gd+ lesions per scan between the BCD-132, 125 mg group and the teriflunomide comparator was [-0,6657 ; - 0,1980], that between the BCD- 132, 500 mg group and teriflunomide group was [-0,6525 ; -0,1897], The results in the groups of the trial BCD- 132 indicate that the upper limit of the confidence interval in both groups does not exceed the preset limit of non-inferiority 0,9975 (6=0,9975).

When comparing the arithmetic means of total T1 Gd+ lesions per scan between the BCD- 132, 125 mg and teriflunomide groups, as well as BCD-132, 500 mg and teriflunomide groups, we observed a statistically significant difference (p=0.0004 - for comparing the BCD-132, 125 mg group and teriflunomide, p=0.0005 - for comparing the BCD-132, 500 mg group and teriflunomide group, two-sided Student-Welch t-test). Due to the fact that the calculated upper limit of the confidence interval for both BCD- 132 groups does not exceed the preset limit of non-inferior efficacy 0.9975 (6=0.9975), the hypothesis of non-inferior efficacy of the trial groups over the teriflunomide comparator at the primary endpoint has been proven.

This indicates the efficacy of therapy and the achievement of the main goal of the trial.

Dynamic estimation of means of total accumulating gadolinium (Gd+) lesions in T1 mode (there are two main types of MRI images: T1 -weighted image and T2-weighted image. The Tl- weighted image registers the rate at which atoms return from the lateral position to the vertical position. The T2-weighted image, the time of retention of atoms in the lateral position following a short-term perpendicular pulse, showed decreased lesions in the divozilimab and teriflunomide groups: in the teriflunomide group, the dynamics was not statistically significant; in the divozilimab groups, a statistically significant decrease relative to screening was indicated at week 9 already.

Assessment of the complex CUA (cumulative new T1 contrast-enhancing MRI lesions and new T2 lesions mode or cases of T2 enlarging lesions, avoiding double counting) using per scan analysis showed that the means for the estimated period were statistically significantly lower in the divozilimab groups as compared to the teriflunomide and placebo groups (p <0.05). Dynamic observation of CUA showed a significant decrease in the means of the indicator in both divozilimab groups (p<0.05; non-parametric Wilcoxon test with Benjamini-Yekutieli adjustment). There was no significant dynamics in the teriflunomide and placebo groups.

A comparable pattern in the form of a decreased demyelinating process activity was also observed during assessment among patients without lesions. Most patients in the divozilimab groups had no T1 contrast-enhancing lesions following 24 weeks of treatment (94.44% and 93.06%). These values were statistically significantly higher than those in the teriflunomide and placebo groups (68.06 and 56.36%) (p<0.05)).

Assessment of other endpoints represented by MRI metrics showed a statistically significant decrease in the volume of hypointensive lesions and the number of new/enlarged T2WI/FLAIR lesions over a period of 24 weeks, including when comparing among patients with new or enlarged lesions in the divozilimab groups. The median annual exacerbation frequency (ARR) in the trial groups calculated using Poisson regression was lower in the teriflunomide and divozilimab (125 and 500) groups. The ARR index was statistically significantly lower in the divozilimab 500mg group.

The EDSS (extended disability status scale) score showed positive dynamics of the patients' condition, revealing a statistically significant decrease in the score following 1 cycle of therapy in the divozilimab 500 mg group. Total proportion of patients who had persistent progression following 24 weeks of therapy was 4.17% in the teriflunomide group and in the BCD-132, 125 mg group, 1.39% in the BCD-132, 500 mg group and 7.27% in the placebo group (p=0.3999; Fisher's exact test). When assessing neurological deficits and cognitive functions using a number of other scales and questionnaires (Timed 7.62 meter/25-foot walk test, 9-hole peg test (9-NHT), symbol digit modalities test (SDMT)), as well as when assessing the quality of patients' life (SF-36, EQ-5D, MSQOL-54), there was no significant negative dynamics in the groups.

The main efficacy results following 24 weeks of therapy in the BCD-132-2 trial are shown in Table 12.

Table 12 - Main efficacy parameters following 24 weeks of therapy in the BCD-132-2 trial

The proportion of patients without confirmed exacerbations is shown in Table 13. In the BCD-132 groups, it was 93.06% and 97.22% of patients (125 mg and 500 mg, respectively) by week 24 of therapy. All patients with exacerbations underwent a brain MRI examination with contrast prior to glucocorticosteroids (GCs) therapy. If MRI results revealed contrast-enhancing T1 lesions and new and/or enlarged T2-WI and T2-FLAIR lesions, then such an exacerbation was considered an MRI-confirmed exacerbation. Such exacerbations were absent in the divozilimab groups by week 24 of therapy, whereas the proportion was 5.56% and 7.27% in the teriflunomide and placebo groups, respectively. The proportion of patients with AINC (Assessing Independent Neurological Commission)-confirmed exacerbations was 6.94% and 1.39% in the BCD-132 groups (125 mg and 500 mg, respectively) as compared to 11.11% and 20.00% in the TFL and PBO groups, respectively.

Table 13 - Main efficacy parameters associated with exacerbations following 24 weeks of therapy within the BCD-132-2 trial

Evaluation of the dynamics of individual key parameters of brain MRI and clinical indicators demonstrated a positive therapy effect: a decreased activity of the demyelinating process in the brain and a decreased exacerbations in patients. A slightly more pronounced effect was observed in the arm that received a higher trial divozilimab dosage compared to a lower dosage, with comparable safety parameters.

The resulting data support efficacy of divozilimab in the treatment of patients with multiple sclerosis with exacerbations.

Clinical safety results summary from trial No. BCD- 132-2.

Throughout the assessed period, all trial arms demonstrated an acceptable safety profile. The therapy carried out in the target population of patients with multiple sclerosis with exacerbations was tolerated satisfactorily. The analysis using the safety endpoints did not reveal any statistically significant differences. According to the investigators, the AEs associated with trial intravenous medicinal product, in organs and systems during the analyzed period, were mostly (more than 5% of patients) respiratory tract infections, infusion reactions, deviations in laboratory and instrumental data (decreased neutrophils, decreased lymphocytes, decreased leukocytes). Inter alia, we also observed infectious conditions such as: infection caused by herpes virus and oral herpes, infection caused by fungi of the genus Candida and enterovirus infection, urinary tract infection and cystitis, pneumonia.

Deviations of other organs and systems were observed in less than 5% of patients. The registered AEs mostly had a mild and moderate degree (grade 1-2). According to the investigators, there were no statistically significant differences between the arms in any of the deviations associated with the trial intravenous medicinal product. AEs of grades 3-4 according to CTCAE v. 5.0, which were associated, according to the investigator, with intravenous medicinal product were represented by deviations of laboratory and instrumental data, liver/ biliary tract disorders and vascular disorders.

Conclusion The study was conducted on a representative population, which was balanced by the main demographic parameters and characteristics of the disease, and which was close to the target population of patients with multiple sclerosis.

The resulting efficacy and safety data justify the use of divozilimab at doses of 125 and 500 mg.

Multicenter open-label phase II trial No. BCD-132-2 in a population of patients with multiple sclerosis demonstrated a significant therapeutic effect of the use of divozilimab. Taking into account the efficacy indicators under a satisfactory safety profile obtained in the trial, the use of divozilimab provides for the risk-benefit balance in the given population of patients.

Claims

1. A pharmaceutical composition of anti-CD20 antibody, comprising:

(i) anti-CD20 antibody;

(ii) acetate buffer;

(iii) sorbitol;

(iv) methionine;

(v) water for inj ection.

The pharmaceutical composition according to claim 1, wherein the anti-CD20 antibody is present at a concentration of 1.5-60.0 MF/MJI.

3. The pharmaceutical composition according to claim 1, wherein the anti-CD20 antibody is present at a concentration of 5.0-50.0 MF/MJI.

4. The pharmaceutical composition according to claim 1, wherein the anti-CD20 antibody is present at a concentration of 10.0-40.0 MF/MJI.

5. The pharmaceutical composition according to claim 1, wherein the anti-CD20 antibody is present at a concentration of 20.0-30.0 MF/MJI.

6. The pharmaceutical composition according to claim 1, wherein the anti-CD20 antibody is present at a concentration of 25.0 MF/MJI.

7. The pharmaceutical composition according to claim 1, wherein the anti-CD20 antibody is divozilimab.

8. The pharmaceutical composition according to claim 1, wherein the acetate buffer is a mixture of sodium acetate and acetic acid.

9. The pharmaceutical composition according to claim 8, wherein sodium acetate is present at a concentration of 0.5-3.0 mg/ml.

10. The pharmaceutical composition according to claim 8, wherein sodium acetate is present at a concentration of 1.0-2.0 mg/ml.

11. The pharmaceutical composition according to claim 8, wherein sodium acetate is present at a concentration of 1.5-2.0 mg/ml.

12. The pharmaceutical composition according to claim 8, wherein sodium acetate is present at a concentration of 1.74 mg/ml.

13. The pharmaceutical composition according to claim 8, wherein sodium acetate is sodium acetate trihydrate.

14. The pharmaceutical composition according to claim 8, wherein acetic acid is added to pH 5.0-6.0.

15. The pharmaceutical composition according to claim 8, wherein acetic acid is added to pH 5.4-5.6.

16. The pharmaceutical composition according to claim 8, wherein acetic acid is added to pH 5.5.

17. The pharmaceutical composition according to claim 8, wherein acetic acid is glacial acetic acid.

18. The pharmaceutical composition according to claim 1, wherein sorbitol is present at a concentration of 40.0-60.0 mg/ml.

19. The pharmaceutical composition according to claim 1, wherein sorbitol is present at a concentration of 45.0-55.0 mg/ml.

20. The pharmaceutical composition according to claim 1, wherein sorbitol is present at a concentration of 50.0 mg/ml.

21. The pharmaceutical composition according to claim 1, wherein methionine is present at a concentration of 0.05-0.30 mg/ml.

22. The pharmaceutical composition according to claim 1, wherein methionine is present at a concentration of 0.10-0.20 mg/ml.

23. The pharmaceutical composition according to claim 1, wherein methionine is present at a concentration of 0.15 mg/ml.

24. The pharmaceutical composition according to claim 1, comprising:

(i) anti-CD20 antibody; acetate buffer being a mixture of sodium acetate 0.5-3.0 mg/ml and acetic acid to pH 5.0-6.0; sorbitol 40.0-60.0 mg/ml;

(iv) methionine 0.05-0.30 mg/ml;

(v) water for injection to 1 ml. 25. The pharmaceutical composition according to claim 1, comprising:

(i) anti-CD20 antibody; acetate buffer being a mixture of sodium acetate 1.0-2.0 mg/ml and acetic acid to pH 5.4-5.6; sorbitol 45.0-55.0 mg/ml;

(iv) methionine 0.10-0.20 mg/ml;

(v) water for injection to 1 ml.

26. The pharmaceutical composition according to claim 1, comprising: anti-CD20 antibody; acetate buffer being a mixture of sodium acetate 1.5-2.0 mg/ml and acetic acid to pH 5.4-5.6;

(vi) sorbitol 45.0-55.0 mg/ml;

(vii) methionine 0.10-0.20 mg/ml;

(viii) water for injection to 1 ml.

27. The pharmaceutical composition according to claim 1, comprising:

(i) anti-CD20 antibody; acetate buffer being a mixture of sodium acetate 1.74 mg/ml and acetic acid to pH 5.5; sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for injection to 1 ml. 28. The pharmaceutical composition according to claim 1, comprising:

(i) anti-CD20 antibody; acetate buffer being a mixture of sodium acetate trihydrate 1.74 mg/ml and acetic acid to pH 5.5; sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for injection to 1 ml. 29. The pharmaceutical composition according to claim 1, comprising:

(i) anti-CD20 antibody 25.0 mg/ml; acetate buffer being a mixture of sodium acetate 1.74 mg/ml and acetic acid to pH 5.5; sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for injection to 1 ml. 30. The pharmaceutical composition according to claim 1, comprising:

(i) divozilimab 25.0 mg/ml; acetate buffer being a mixture of sodium acetate 1.74 mg/ml and acetic acid to pH 5.5; sorbitol 50.0 mg/ml; methionine 0.15 mg/ml; water for injection to 1 ml.

The pharmaceutical composition according to claim 1, comprising: anti-CD20 antibody 25.0 mg/ml; acetate buffer being a mixture of sodium acetate trihydrate 1.74 mg/ml and acetic acid to pH 5.5; sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for injection to 1 ml. 32. The pharmaceutical composition according to claim 1, comprising:

(i) divozilimab 25.0 mg/ml; acetate buffer being a mixture of sodium acetate trihydrate 1.74 mg/ml and acetic acid to pH 5.5; sorbitol 50.0 mg/ml;

(iv) methionine 0.15 mg/ml;

(v) water for injection to 1 ml.

33. Use of the pharmaceutical composition according to any one of claims 1-32 for the treatment of a CD20-mediated disease or disorder in a subject in need thereof.

34. The use according to claim 33, wherein the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 125 mg.

35. The use according to claim 33, wherein the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 500 mg.

36. The use according to claim 33, wherein the pharmaceutical composition is administered every 24 weeks.

37. The use according to claim 33, wherein the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 125 mg every 24 weeks.

38. The use according to claim 33, wherein the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 500 mg every 24 weeks.

39. The use according to claim 33, comprising at least 5 cycles of administration of the pharmaceutical composition.

40. The use according to claim 33, comprising at least 5 cycles of administration of the pharmaceutical composition, wherein cycle 1 includes the first administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 62.5 mg and the second administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 62.5 mg 2 weeks following the first administration; the following cycles include the administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 125 mg every 24 weeks following the first administration.

41. The use according to claim 33, comprising at least 5 cycles of administration of the pharmaceutical composition, wherein cycle 1 includes the first administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 250 mg and the second administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 250 mg 2 weeks following the first administration; the following cycles include the administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 500 mg every 24 weeks following the first administration.

42. The use according to claim 33, wherein the anti-CD20 antibody is divozilimab.

43. The use according to claim 33, wherein the CD20-mediated disease or disorder is selected from the group:

(i) oncological disease or disorder or

(ii) autoimmune disease or disorder.

44. The use according to claim 43, wherein the oncological disease or disorder is selected from the group: B-cell lymphoma, leukemia.

45. The use according to claim 44, wherein B-cell lymphoma is non-Hodgkin's lymphoma, Hodgkin's lymphoma.

46. The use according to claim 44, wherein leukemia is selected from the group: chronic lymphocytic leukemia, small lymphocytic lymphoma.

47. The use according to claim 43, wherein the autoimmune disease or disorder is selected from the group: multiple sclerosis, axonal neuropathy, ANCA-associated vasculitides, IgG4-related disease, amyloidosis, axial spondyloarthritis, antiphospholipid syndrome, Takayasu's arteritis, autoimmune aplastic anemia, autoimmune inner ear disease, autoimmune hemolytic anemia, autoimmune hyperlipidemia, autoimmune dysautonomia, autoimmune urticaria, autoimmune neuromyotonia (Isaacs syndrome), autoimmune retinopathy, autoimmune thrombocytopenic purpura, warm antibody hemolytic anemia, autoimmune testicular disease, autoimmune angioedema, autoimmune hepatitis, autoimmune diabetes, autoimmune immunodeficiency, autoimmune pericarditis, autoimmune thyroiditis, autoimmune progesterone dermatitis, Addison's disease, Behcet's disease, Graves' disease (diffuse toxic goiter), neuromyelitis optica spectrum diseases (Devic's disease), Castleman's disease, Crohn's disease, Ormond's disease (retroperitoneal fibrosis), Raynaud's disease, Churg-Strauss syndrome, bullous pemphigoid, epidermolysis bullosa, vasculitis, renal vasculitis, antineutrophil cytoplasmic antibody-associated vasculitides, vesiculobullous dermatosis, giant cell arteritis, vitiligo, lupus nephritis, inflammatory aortic aneurysm, inflammatory bowel disease, congenital heart block, pemphigus vulgaris, dermatitis herpetiformis, gestational pemphigoid, hypogammaglobulinemia, glomerulonephritis, granulomatosis with polyangiitis (Wegener's syndrome), dermatomyositis, dilated cardiomyopathy, demyelinating neuropathy, discoid lupus erythematosus, idiopathic recurrent macrohematuria (IgA nephropathy), idiopathic inflammatory myopathy, idiopathic inflammatory pseudotumor, idiopathic thrombocytopenic purpura, idiopathic hypocomplementemic tubulointerstitial nephritis, idiopathic pulmonary fibrosis, diabetes mellitus, pure red cell aplasia, Balo's concentric sclerosis, lichen planus, lichen sclerosus, leukocytoclastic vasculitis, limited scleroderma (CREST syndrome), limbic encephalitis, linear IgA bullous dermatosis, pemphigus foliaceus, Waldenstrom macroglobulinemia, mediastinal fibrosis, myasthenic crisis, Lambert-Eaton myasthenic syndrome, myasthenia gravis, microscopic polyangiitis, myositis, Coxsackie myocarditis, multiple sclerosis, multifocal motor neuropathy, multifocal fibrosclerosis, optic neuritis, undifferentiated connective tissue disease uveitis, neutropenia, ulcerative colitis, anti-tubular/glomerular basement membrane antibody-associated nephritis, opsoclonus, acute motor axonal neuropathy, acute hemorrhagic leukoencephalitis, acute disseminated encephalomyelitis, palindromic rheumatism, paraneoplastic cerebellar degeneration, paraneoplastic syndrome, paraproteinemic polyneuropathy, paroxysmal nocturnal hemoglobinuria, ocular pemphigus, mucous membrane pemphigoid, primary biliary cirrhosis, primary sclerosing cholangitis, periaortitis, periarteritis, perivenous encephalomyelitis, peripheral neuropathy, peripheral uveitis, pernicious anemia, subacute bacterial endocarditis, polymyositis, polyneuropathy with organomegaly (POEMS syndrome), transverse myelitis, Dressier's syndrome, post-traumatic pericarditis, psoriasis, psoriatic arthritis, pemphigus, Henoch-Schonlein purpura, anti-HLA antibody-mediated transplant rejection, reactive arthritis, rheumatic fever, polymyalgia rheumatica, rheumatoid arthritis, reflex sympathetic dystrophy, refractory epilepsy, relapsing polychondritis, sarcoidosis, Guillain-Barre syndrome, Kawasaki syndrome, Cogan syndrome, Mikulicz syndrome, Stiff-person syndrome, Sjogren's syndrome, Evans syndrome, systemic lupus erythematosus, systemic scleroderma, limited scleroderma, Riedel's thyroiditis, Hashimoto's thyroiditis, thyroid-associated ophthalmopathy, thyroid crisis, thrombotic thrombocytopenic purpura, polyarteritis nodosa, fetal and neonatal alloimmune thrombocytopenia, fibrosing alveolitis, B-cell non-Hodgkin's lymphoma, B-cell chronic lymphocytic leukemia, cold haemagglutinin disease, Sydenham's chorea (chorea minor or rheumatic chorea), chronic Lyme disease, chronic inflammatory demyelinating polyneuropathy, chronic recurrent multifocal osteomyelitis, chronic sclerosing sialadenitis (Kuttner's tumor), celiac disease, endometriosis, encephalitis, autoantibody -associated encephalopathy, Hashimoto's encephalitis, eosinophilic fasciitis, essential mixed cryoglobulinemia, juvenile diabetes, juvenile rheumatoid arthritis.

48. The use according to claim 47, wherein multiple sclerosis is remitting multiple sclerosis, secondary progressive multiple sclerosis, primary progressive multiple sclerosis, multiple sclerosis with exacerbations, highly active multiple sclerosis, aggressive multiple sclerosis, malignant multiple sclerosis (Marburg's variant), myelinoclastic diffuse Schilder's sclerosis, myelocortical multiple sclerosis, a clinically isolated syndrome.

49. A method of treating a CD20-mediated disease or disorder, comprising administering a pharmaceutical composition according to any one of claims 1-30 to a subject in need thereof in a therapeutically effective amount.

50. The method of treating a disease or disorder according to claim 49, wherein the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 125 mg.

51. The method of treating a disease or disorder according to claim 49, wherein the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 500 mg.

52. The method of treating a disease or disorder according to claim 49, wherein the pharmaceutical composition is administered every 24 weeks.

53. The method of treating a disease or disorder according to claim 49, wherein the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 125 mg every 24 weeks.

54. The method of treating a disease or disorder according to claim 49, wherein the pharmaceutical composition is administered at a dose of anti-CD20 antibody of 500 mg every 24 weeks.

55. The method of treating a disease or disorder according to claim 49, comprising at least 5 cycles of administration of the pharmaceutical composition.

56. The method of treating a disease or disorder according to claim 49, comprising at least 5 cycles, wherein cycle 1 includes the first administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 62.5 mg and the second administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 62.5 mg 2 weeks following the first administration; the following cycles include the administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 125 mg every 24 weeks following the first administration.

57. The method of treating a disease or disorder according to claim 49, comprising at least 5 cycles, wherein cycle 1 includes the first administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 250 mg and the second administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 250 mg 2 weeks following the first administration; the following cycles include the administration of the pharmaceutical composition at a dose of anti-CD20 antibody of 500 mg every 24 weeks following the first administration.

58. The method of treating a disease or disorder according to claim 49, wherein the anti-CD20 antibody is divozilimab.

59. The method of treating a disease or disorder according to claim 49, wherein the CD20-mediated disease or disorder is selected from the group:

(i) oncological disease or disorder or

(ii) autoimmune disease or disorder.

60. The method of treating a disease or disorder according to claim 59, wherein the oncological disease or disorder is selected from the group: B-cell lymphoma, leukemia.

61. The method of treating a disease or disorder according to claim 60, wherein B-cell lymphoma is non-Hodgkin's lymphoma, Hodgkin's lymphoma.

62. The method of treating a disease or disorder according to claim 60, wherein leukemia is selected from the group: chronic lymphocytic leukemia, small lymphocytic lymphoma.

63. The method of treating a disease or disorder according to claim 59, wherein the autoimmune disease or disorder is selected from the group: multiple sclerosis, axonal neuropathy, ANCA-associated vasculitides, IgG4-related disease, amyloidosis, axial spondyloarthritis, antiphospholipid syndrome, Takayasu's arteritis, autoimmune aplastic anemia, autoimmune inner ear disease, autoimmune hemolytic anemia, autoimmune hyperlipidemia, autoimmune dysautonomia, autoimmune urticaria, autoimmune neuromyotonia (Isaacs syndrome), autoimmune retinopathy, autoimmune thrombocytopenic purpura, warm antibody hemolytic anemia, autoimmune testicular disease, autoimmune angioedema, autoimmune hepatitis, autoimmune diabetes, autoimmune immunodeficiency, autoimmune pericarditis, autoimmune thyroiditis, autoimmune progesterone dermatitis, Addison's disease, Behcet's disease, Graves' disease (diffuse toxic goiter), neuromyelitis optica spectrum diseases (Devic's disease), Castleman's disease, Crohn's disease, Ormond's disease (retroperitoneal fibrosis), Raynaud's disease, Churg-Strauss syndrome, bullous pemphigoid, epidermolysis bullosa, vasculitis, renal vasculitis, antineutrophil cytoplasmic antibody-associated vasculitides, vesiculobullous dermatosis, giant cell arteritis, vitiligo, lupus nephritis, inflammatory aortic aneurysm, inflammatory bowel disease, congenital heart block, pemphigus vulgaris, dermatitis herpetiformis, gestational pemphigoid, hypogammaglobulinemia, glomerulonephritis, granulomatosis with polyangiitis (Wegener's syndrome), dermatomyositis, dilated cardiomyopathy, demyelinating neuropathy, discoid lupus erythematosus, idiopathic recurrent macrohematuria (IgA nephropathy), idiopathic inflammatory myopathy, idiopathic inflammatory pseudotumor, idiopathic thrombocytopenic purpura, idiopathic hypocomplementemic tubulointerstitial nephritis, idiopathic pulmonary fibrosis, diabetes mellitus, pure red cell aplasia, Balo's concentric sclerosis, lichen planus, lichen sclerosus, leukocytoclastic vasculitis, limited scleroderma (CREST syndrome), limbic encephalitis, linear IgA bullous dermatosis, pemphigus foliaceus, Waldenstrom macroglobulinemia, mediastinal fibrosis, myasthenic crisis, Lambert-Eaton myasthenic syndrome, myasthenia gravis, microscopic polyangiitis, myositis, Coxsackie myocarditis, multiple sclerosis, multifocal motor neuropathy, multifocal fibrosclerosis, optic neuritis, undifferentiated connective tissue disease uveitis, neutropenia, ulcerative colitis, anti-tubular/glomerular basement membrane antibody-associated nephritis, opsoclonus, acute motor axonal neuropathy, acute hemorrhagic leukoencephalitis, acute disseminated encephalomyelitis, palindromic rheumatism, paraneoplastic cerebellar degeneration, paraneoplastic syndrome, paraproteinemic polyneuropathy, paroxysmal nocturnal hemoglobinuria, ocular pemphigus, mucous membrane pemphigoid, primary biliary cirrhosis, primary sclerosing cholangitis, periaortitis, periarteritis, perivenous encephalomyelitis, peripheral neuropathy, peripheral uveitis, pernicious anemia, subacute bacterial endocarditis, polymyositis, polyneuropathy with organomegaly (POEMS syndrome), transverse myelitis, Dressier's syndrome, post-traumatic pericarditis, psoriasis, psoriatic arthritis, pemphigus, Henoch- Schonlein purpura, anti-HLA antibody-mediated transplant rejection, reactive arthritis, rheumatic fever, polymyalgia rheumatica, rheumatoid arthritis, reflex sympathetic dystrophy, refractory epilepsy, relapsing polychondritis, sarcoidosis, Guillain-Barre syndrome, Kawasaki syndrome, Cogan syndrome, Mikulicz syndrome, Stiff-person syndrome, Sjogren's syndrome, Evans syndrome, systemic lupus erythematosus, systemic scleroderma, limited scleroderma, Riedel's thyroiditis, Hashimoto's thyroiditis, thyroid-associated ophthalmopathy, thyroid crisis, thrombotic thrombocytopenic purpura, polyarteritis nodosa, fetal and neonatal alloimmune thrombocytopenia, fibrosing alveolitis, B-cell non-Hodgkin's lymphoma, B-cell chronic lymphocytic leukemia, cold haemagglutinin disease, Sydenham's chorea (chorea minor or rheumatic chorea), chronic Lyme disease, chronic inflammatory demyelinating polyneuropathy, chronic recurrent multifocal osteomyelitis, chronic sclerosing sialadenitis (Kiittner's tumor), celiac disease, endometriosis, encephalitis, autoantibody -associated encephalopathy, Hashimoto's encephalitis, eosinophilic fasciitis, essential mixed cryoglobulinemia, juvenile diabetes, juvenile rheumatoid arthritis.

64. The method of treating a disease or disorder according to claim 63, wherein multiple sclerosis is remitting multiple sclerosis, secondary progressive multiple sclerosis, primary progressive multiple sclerosis, multiple sclerosis with exacerbations, highly active multiple sclerosis, aggressive multiple sclerosis, malignant multiple sclerosis (Marburg's variant), myelinoclastic diffuse Schilder's sclerosis, myelocortical multiple sclerosis, a clinically isolated syndrome.