Aqueous pharmaceutical compositions of anti-pd-1 antibody prolglolimab and uses thereof

Abstract

The present application relates to an aqueous pharmaceutical composition of anti-PD-1 antibody PROLGOLIMAB and its application, the aqueous pharmaceutical composition contains a pharmaceutically effective amount of anti-PD-1 antibody Prolgolimab, an effective amount of trehalose dihydrate, an acetate or histidine-based buffer agent.Experiments have proved that the aqueous pharmaceutical composition of the present application has higher aggregation stability and affinity than the known anti-PD-1 antibody based on IgG4 isotype human antibody, and can be used for preparing a pharmaceutical preparation for treating malignant tumors.

Description

Technical Field

[0001] This invention relates to novel aqueous compositions of anti-PD-1 antibodies, and more particularly to novel aqueous compositions of the anti-PD-1 antibody Prolgolimab, which can be used as a medicament for treating malignant tumors. Background Technology

[0002] Programmed death 1 (PD-1) is an inhibitory protein of the CD28 receptor family, which, in addition to PD-1, also includes CD28, CTLA-4, ICOS, and BTLA. PD-1 is expressed by activated B cells, T cells, and myeloid cells (Agata et al., supra; Okazaki et al. (2002) Curr. Opin. Immunol. 14:391779-82; Bennet et al. (2003) J Immunol 170:711-8). As initial members of this receptor family, CD28 and ICOS were discovered for their role in promoting T cell proliferation upon the addition of monoclonal antibodies (Hutloff et al. (1999) Nature 397:263-266; Hansen et al. (1980) Immunogenics 10:247-260). PD-1 was identified in differential expression screening studies in apoptotic cells (Ishida et al. (1992) EMBO J 11:3887-95). Other members of the receptor family—CTLA-4 and BTLA—were identified in differential expression screening studies in cytotoxic T lymphocytes and TH1 cells, respectively. CD28, ICOS, and CTLA-4 all possess unpaired cysteine ​​residues that enable their homodimerization. In contrast, PD-1 is believed to exist in monomeric form and lacks the unpaired cysteine ​​residue characteristic of other members of the CD28 family.

[0003] PD-1 is a 55 kDa type I transmembrane protein and a member of the Ig gene superfamily (Agata et al. (1996) Int Immunol 8:765-72). PD-1 contains a proximal immunoreceptor tyrosine repressor motif (ITIM) and a distal immunoreceptor tyrosine switching motif (ITSM) (Thomas, ML (1995) J Exp Med 181:1953-6; Vivier, E. Daeron, M (1997) Immunol Today 18:286-91). Although PD-1 is structurally similar to CTLA-4, it lacks the MYPPY motif required for the binding of B7-1 and B7-2. It has been found that PD-1 has two ligands—PD-L1 and PD-L2—and these two ligands have been shown to have a negative regulatory effect on T cell activation after binding to PD-1 (Freeman et al. (2000) J Exp Med 192:1027-34; Latchman et al. (2001) Nat Immunol 2:261-8; Carter et al. (2002) Eur J Immunol 32:634-43). Both PD-L1 and PD-L2 are B7 homologs that can bind to PD-1 but not to other members of the CD28 family.

[0004] As a PD-1 ligand, PD-L1 is abundant in various types of human cancers (Dong et al. (2002) Nat. Med. 8:787-9). The interaction between PD-1 and PD-L1 reduces the number of tumor-infiltrating lymphocytes, decreases T cell receptor-mediated proliferation, and allows cancer cells to escape immune surveillance mechanisms (Dong et al. (2003) J. Mol. Med. 81:281-7; Blank et al. (2005) Cancer Immunol. Immunother. 54:307-314; Konishi et al. (2004) Clin. Cancer Res. 10:5094-100). Immunosuppressive mechanisms can be reversed by inhibiting local PD-L1 / PD-1 interactions, and this reversal is additive when PD-L2 / PD-1 interactions are blocked (Iwai et al. (2002) Proc. Nat'l. Acad. Sci. USA 99:12293-7; Brown et al. (2003) J. Immunol. 170:1257-66).

[0005] PD-1 is a repressive member of the CD28 family and is expressed by activated B cells, T cells, and myeloid cells (Agata et al., supra; Okazaki et al. (2002) Curr Opin Immunol 14:391779-82; Bennett et al. (2003) J Immunol 170:711-8). Animals lacking PD-1 are prone to developing various autoimmune diseases, including autoimmune heart disease and lupus-like syndromes including arthritis and nephritis (Nishimura et al. (1999) Immunity 11:141-51; Nishimura et al. (2001) Science 291:319-22). Furthermore, PD-1 has been found to have certain effects on autoimmune encephalomyelitis, systemic lupus erythematosus, graft-versus-host disease (GVHD), type 1 diabetes, and rheumatoid arthritis (Salama et al. (2003) J Exp Med 198:71-78; Prokunina and Alarcon-Riquelme (2004) Hum Mol Genet 13:R143; Nielsene et al. (2004) Lupus 13:510). In mouse B-cell tumor lines, the ITSM of PD-1 has been shown to block the downstream effector molecule Ca2+ mediated by BCR. 2+ Essential for channels and tyrosine phosphorylation (Okazaki et al. (2001) PNAS 98:13866-71).

[0006] Several known anti-PD-1 antibodies exist in this field, such as nivolumab (BMS) and pembrolizumab (Merck), which are human IgG4 monoclonal antibodies.

[0007] Another known anti-PD-1 antibody is the novel antibody Prolgolimab (also known as BCD-100), an IgG1 isotype monoclonal human antibody with non-effect mutants L234A and L235A. It has been shown that Prolgolimab has a higher affinity for PD-1 and greater aggregation stability compared to IgG4 antibodies. Furthermore, Prolgolimab is currently in clinical trials for various types of malignancies, including: melanoma, including inoperable or metastatic melanoma, and early-stage melanoma before and after definitive treatment; and lung cancer, specifically non-small cell lung cancer (NSCLC), including inoperable or metastatic NSCLC.

[0008] Based on the above, it is clear that creating a new and improved stable aqueous drug composition for the anti-PD-1 antibody Prolgolimab is of great significance. Summary of the Invention

[0009] This invention discloses an aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab, which has higher aggregation stability and affinity than known anti-PD-1 antibodies based on IgG4 isotype human antibodies.

[0010] As disclosed in International Patent Application WO / 2018 / 013017 (incorporated herein by reference), the anti-PD-1 antibody Prolgolimab is an IgG1 isotype monoclonal human antibody with non-effect mutants L234A and L235A (referred to herein as the "antibody of this invention"). Compared with known anti-PD-1 antibodies based on IgG4 isotype human antibodies, such as nivolumab, it exhibits higher aggregation stability, greater affinity, and better t-cell aggregation. 1 / 2β (hours) or C max Pharmacokinetic parameters such as (μg / mL) are available. Prolgolimab has a weight-average molecular weight of approximately 146 kDa and is specific for human PD-1. Prolgolimab has a heavy chain containing 459 amino acids (SEQ ID NO: 1) and a human light chain containing 214 amino acids (SEQ ID NO: 2). The constant moiety (Fc) of Prolgolimab contains L234A and L235A mutations.

[0011] Therefore, administering an aqueous composition containing the antibody of the present invention to patients with malignant tumors can be beneficial.

[0012] In one general aspect, the present invention is an aqueous pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity. An aqueous pharmaceutical composition contains a pharmaceutically effective amount of the anti-PD-1 antibody Prolgolimab, an effective amount of trehalose dihydrate, and an acetate- or histidine-based buffer.

[0013] According to one general aspect of the present invention, an aqueous pharmaceutical composition for an anti-PD-1 antibody is provided, comprising:

[0014] (a) Prolgolimab at a concentration of 15 mg / mL to 40 mg / mL as an antibody;

[0015] (b) Trehalose dihydrate at a concentration of 80 mg / mL to 110 mg / mL;

[0016] (c) Sodium acetate trihydrate at a concentration of 0.2 mg / mL to 2.5 mg / mL; and

[0017] (d) Add acetic acid to a pH of 4.5–5.5.

[0018] In some embodiments of the present invention, the concentration of Prolgolimab can be 15 mg / mL to 25 mg / mL.

[0019] In some embodiments of the present invention, the concentration of Prolgolimab can be 20 mg / mL.

[0020] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 95 mg / mL to 105 mg / mL.

[0021] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 100 mg / mL.

[0022] In some embodiments of the present invention, the concentration of sodium acetate trihydrate can be 1.6 mg / mL to 1.9 mg / mL.

[0023] In some embodiments of the present invention, the concentration of sodium acetate trihydrate can be 1.7 mg / mL to 1.8 mg / mL.

[0024] In some embodiments of the present invention, the concentration of sodium acetate trihydrate may be 1.742 mg / mL.

[0025] In some embodiments of the present invention, acetic acid may be added to a pH value of 5.0.

[0026] In some embodiments of the present invention, the concentration of acetic acid may be 0.04 mg / mL to 0.77 mg / mL.

[0027] In some embodiments of the present invention, the concentration of acetic acid may be 0.40 mg / mL to 0.50 mg / mL.

[0028] In some embodiments of the present invention, the concentration of acetic acid may be 0.43 mg / mL.

[0029] According to one general aspect of the present invention, an aqueous pharmaceutical composition for an anti-PD-1 antibody is provided, comprising:

[0030] (a) Prolgolimab at a concentration of 90 mg / mL to 150 mg / mL as an antibody;

[0031] (b) Trehalose dihydrate at a concentration of 50 mg / mL to 110 mg / mL;

[0032] (c) Sodium acetate trihydrate at a concentration of 0.2 mg / mL to 2.5 mg / mL; and

[0033] (d) Add acetic acid to a pH of 4.5–5.5.

[0034] In some embodiments of the present invention, the concentration of Prolgolimab can be 90 mg / mL to 110 mg / mL.

[0035] In some embodiments of the present invention, the concentration of Prolgolimab can be 100 mg / mL.

[0036] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 75 mg / mL to 85 mg / mL.

[0037] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 80 mg / mL.

[0038] In some embodiments of the present invention, the concentration of sodium acetate trihydrate can be 1.6 mg / mL to 1.9 mg / mL.

[0039] In some embodiments of the present invention, the concentration of sodium acetate trihydrate can be 1.7 mg / mL to 1.8 mg / mL.

[0040] In some embodiments of the present invention, the concentration of sodium acetate trihydrate may be 1.742 mg / mL.

[0041] In some embodiments of the present invention, acetic acid may be added to a pH value of 5.0 to 5.5.

[0042] In some embodiments of the present invention, acetic acid may be added to a pH value of 5.0.

[0043] In some embodiments of the present invention, the concentration of acetic acid may be 0.045 mg / mL to 0.77 mg / mL.

[0044] In some embodiments of the present invention, the concentration of acetic acid may be 0.40 mg / mL to 0.50 mg / mL.

[0045] In some embodiments of the present invention, the concentration of acetic acid may be 0.43 mg / mL.

[0046] According to one general aspect of the present invention, an aqueous pharmaceutical composition for an anti-PD-1 antibody is provided, comprising:

[0047] (a) Prolgolimab as an antibody at a concentration of 5 mg / mL to 150 mg / mL;

[0048] (b) Trehalose dihydrate at a concentration of 70 mg / mL to 110 mg / mL;

[0049] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0050] (d) L-histidine hydrochloride at a concentration of 0.2–3.5 mg / mL.

[0051] In some embodiments of the present invention, the concentration of Prolgolimab can be from 15 mg / mL to 40 mg / mL.

[0052] In some embodiments of the present invention, the concentration of Prolgolimab can be 15 mg / mL to 25 mg / mL.

[0053] In some embodiments of the present invention, the concentration of Prolgolimab can be 20 mg / mL.

[0054] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 95 mg / mL to 105 mg / mL.

[0055] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 100 mg / mL.

[0056] In some embodiments of the present invention, the concentration of L-histidine can be 0.7 mg / mL to 1.0 mg / mL.

[0057] In some embodiments of the present invention, the concentration of L-histidine may be 0.92 mg / mL.

[0058] In some embodiments of the present invention, the concentration of L-histidine hydrochloride may be 2.8 mg / mL to 3.3 mg / mL.

[0059] In some embodiments of the present invention, the concentration of L-histidine hydrochloride may be 2.96 mg / mL.

[0060] In some embodiments of the present invention, the pH value of the composition may be 5.5 to 6.5.

[0061] In some embodiments of the present invention, the pH value of the composition may be 5.5.

[0062] In some embodiments of the present invention, the concentration of Prolgolimab can be 90 mg / mL to 110 mg / mL.

[0063] In some embodiments of the present invention, the concentration of Prolgolimab can be 100 mg / mL.

[0064] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 75 mg / mL to 85 mg / mL.

[0065] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 80 mg / mL.

[0066] In some embodiments of the present invention, the concentration of L-histidine can be 0.7 mg / mL to 1.0 mg / mL.

[0067] In some embodiments of the present invention, the concentration of L-histidine may be 0.92 mg / mL.

[0068] In some embodiments of the present invention, the concentration of L-histidine hydrochloride may be 2.8 mg / mL to 3.3 mg / mL.

[0069] In some embodiments of the present invention, the concentration of L-histidine hydrochloride may be 2.96 mg / mL.

[0070] In some embodiments of the present invention, the pH value of the composition may be 5.5 to 6.5.

[0071] In some embodiments of the present invention, the pH value of the composition may be 5.5 to 6.0.

[0072] In some embodiments of the present invention, the pH value of the composition may be 5.5.

[0073] An aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention may further contain a suitable solubilizer.

[0074] In some embodiments of the present invention, the solubilizer may be poloxamer 188.

[0075] In some embodiments of the present invention, the content of poloxamer 188 may be greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0076] In some embodiments of the present invention, the content of poloxamer 188 may be 0 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.4 mg / mL, 0.5 mg / mL, 0.6 mg / mL, 0.7 mg / mL, 0.8 mg / mL, 0.9 mg / mL, or 1.0 mg / mL.

[0077] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0078] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0079] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0080] (c) Sodium acetate trihydrate at a concentration of 0.2–2.5 mg / mL; and

[0081] (d) Add acetic acid to a pH of 4.5–5.5.

[0082] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0083] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0084] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0085] (c) Sodium acetate trihydrate at a concentration of 1.7 mg / mL to 1.8 mg / mL; and

[0086] (d) Add acetic acid to a pH of 5.0.

[0087] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0088] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0089] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0090] (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL; and

[0091] (d) Add acetic acid to a pH of 5.0.

[0092] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0093] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0094] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0095] (c) Sodium acetate trihydrate at a concentration of 0.2–2.5 mg / mL; and

[0096] (d) Add acetic acid to a pH of 4.5–5.5.

[0097] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0098] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0099] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0100] (c) Sodium acetate trihydrate at a concentration of 1.7 mg / mL to 1.8 mg / mL; and

[0101] (d) Add acetic acid to a pH of 5.0–5.5.

[0102] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0103] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0104] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0105] (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL; and

[0106] (d) Add acetic acid to a pH of 5.0–5.5.

[0107] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0108] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0109] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0110] (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL; and

[0111] (d) Add acetic acid to a pH of 5.0.

[0112] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0113] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0114] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0115] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0116] (d) L-histidine hydrochloride at a concentration of 0.2–3.5 mg / mL;

[0117] (e) and the pH value of the aqueous pharmaceutical composition is 5.5 to 6.5.

[0118] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0119] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0120] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0121] (c) L-histidine at a concentration of 0.92 mg / mL; and

[0122] (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL;

[0123] (e) and the pH value of the aqueous pharmaceutical composition is 5.5.

[0124] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0125] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0126] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0127] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0128] (d) L-histidine hydrochloride at a concentration of 0.2–3.5 mg / mL;

[0129] (e) and the pH value of the aqueous pharmaceutical composition is 5.5 to 6.5.

[0130] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0131] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0132] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0133] (c) L-histidine at a concentration of 0.92 mg / mL; and

[0134] (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL;

[0135] (e) and the pH value of the aqueous pharmaceutical composition is 5.5 to 6.0.

[0136] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0137] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0138] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0139] (c) L-histidine at a concentration of 0.92 mg / mL; and

[0140] (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL;

[0141] (e) and the pH value of the aqueous pharmaceutical composition is 5.5.

[0142] In one embodiment, the present invention relates to an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0143] I:

[0144]

[0145] In one embodiment, the present invention relates to an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0146] II:

[0147]

[0148] The aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention may further contain a suitable solubilizer.

[0149] In some embodiments of the present invention, the solubilizer may be poloxamer 188.

[0150] In some embodiments of the present invention, the amount of poloxamer 188 may be greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0151] In some embodiments of the present invention, the amount of poloxamer 188 may be 0 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.4 mg / mL, 0.5 mg / mL, 0.6 mg / mL, 0.7 mg / mL, 0.8 mg / mL, 0.9 mg / mL, or 1.0 mg / mL.

[0152] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0153] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0154] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0155] (c) Sodium acetate trihydrate at a concentration of 0.2–2.5 mg / mL;

[0156] (d) Add acetic acid to a pH of 4.5–5.5; and

[0157] (e) Poloxamer 188 at a concentration greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0158] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0159] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0160] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0161] (c) Sodium acetate trihydrate at a concentration of 1.7 mg / mL to 1.8 mg / mL;

[0162] (d) Add acetic acid to a pH of 5.0; and

[0163] (e) Poloxamer 188 at a concentration greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0164] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0165] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0166] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0167] (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL;

[0168] (d) Add acetic acid to a pH of 5.0; and

[0169] (e) Poloxamer 188 at a concentration greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0170] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0171] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0172] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0173] (c) Sodium acetate trihydrate at a concentration of 0.2–2.5 mg / mL;

[0174] (d) Add acetic acid to a pH of 4.5–5.5; and

[0175] (e) Poloxamer 188 at a concentration greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0176] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0177] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0178] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0179] (c) Sodium acetate trihydrate at a concentration of 1.7 mg / mL to 1.8 mg / mL;

[0180] (d) Add acetic acid to a pH of 5.0–5.5; and

[0181] (e) Poloxamer 188 at a concentration greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0182] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0183] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0184] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0185] (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL;

[0186] (d) Add acetic acid to a pH of 5.0–5.5; and

[0187] (e) Poloxamer 188 at a concentration greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0188] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0189] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0190] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0191] (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL;

[0192] (d) Add acetic acid to a pH of 5.0; and

[0193] (e) Poloxamer 188 at a concentration greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0194] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0195] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0196] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0197] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0198] (d) L-histidine hydrochloride at a concentration of 0.2–3.5 mg / mL;

[0199] (e) wherein the pH value of the composition is 5.5 to 6.5.

[0200] (f) The concentration of poloxamer 188 is greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0201] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0202] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0203] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0204] (c) L-histidine at a concentration of 0.92 mg / mL; and

[0205] (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL;

[0206] (e) wherein the pH value of the composition is 5.5,

[0207] (f) The concentration of poloxamer 188 is greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0208] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0209] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0210] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0211] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0212] (d) L-histidine hydrochloride at a concentration of 0.2–3.5 mg / mL;

[0213] (e) wherein the pH value of the composition is 5.5 to 6.5.

[0214] (f) The concentration of poloxamer 188 is greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0215] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0216] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0217] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0218] (c) L-histidine at a concentration of 0.92 mg / mL; and

[0219] (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL;

[0220] (e) wherein the pH value of the composition is 5.5 to 6.0,

[0221] (f) The concentration of poloxamer 188 is greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0222] In one embodiment, the present invention relates to an aqueous pharmaceutical composition containing an anti-PD-1 antibody, comprising:

[0223] (a) Prolgolimab at a concentration of 100 mg / mL as an antibody;

[0224] (b) Trehalose dihydrate at a concentration of 80 mg / mL;

[0225] (c) L-histidine at a concentration of 0.92 mg / mL; and

[0226] (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL;

[0227] (e) wherein the pH value of the composition is 5.5,

[0228] (f) The concentration of poloxamer 188 is greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0229] In some embodiments of the present invention, the amount of poloxamer 188 may be 0 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.4 mg / mL, 0.5 mg / mL, 0.6 mg / mL, 0.7 mg / mL, 0.8 mg / mL, 0.9 mg / mL, or 1.0 mg / mL.

[0230] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody of the present invention can be administered parenterally.

[0231] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody of the present invention can be administered intramuscularly.

[0232] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody of the present invention can be administered subcutaneously.

[0233] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody of the present invention can be administered intravenously.

[0234] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody of the present invention can be administered via intravenous infusion.

[0235] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody of the present invention can be administered by intravenous infusion over a period of 60 minutes. In cases of good tolerability, the infusion time can be shortened to 30 minutes.

[0236] In one embodiment, the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention can be placed in a small vial.

[0237] In some embodiments, the small medicine bottle can be a glass medicine bottle.

[0238] In some embodiments, the volume of the vial can be 1 mL to 50 mL.

[0239] In some embodiments, the volume of the vial can be 1 mL to 20 mL.

[0240] In some embodiments, the volume of the vial can be 1 mL, 2 mL, 3 mL, 4 mL, 5 mL, 6 mL, 7 mL, 8 mL, 9 mL, 10 mL, 15 mL, 20 mL, 25 mL, 30 mL, 35 mL, 40 mL, 45 mL, or 50 mL.

[0241] In one embodiment, the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention can be placed in a syringe.

[0242] In some embodiments, the syringe may have a volume of 1 mL.

[0243] In some embodiments, the syringe may have a volume of 2 mL.

[0244] In one embodiment, the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention can be placed in a pre-filled syringe.

[0245] In some embodiments, the volume of the pre-filled syringe may be 1 mL.

[0246] In some embodiments, the volume of the pre-filled syringe may be 2 mL.

[0247] In another general aspect, the present invention relates to a method for manufacturing an aqueous pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity. The method comprises combining a pharmaceutically effective amount of the anti-PD-1 antibody Prolgolimab with an acetate-based buffer and an effective amount of trehalose. The method further comprises combining a pharmaceutically effective amount of the anti-PD-1 antibody Prolgolimab with a histidine-based buffer and an effective amount of trehalose.

[0248] In some embodiments, poloxamer 188 may be added as a solubilizer.

[0249] In another general aspect, the present invention relates to the use of the PD-1 antibody Prolgolimab in the malignant tumor treatment of the water-resistant pharmaceutical composition described herein.

[0250] In some embodiments of the present invention, the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab is used for the treatment of malignant tumors, which may be selected from the group comprising: melanoma, including inoperable melanoma or metastatic melanoma, and early melanoma before and after definitive treatment; lung cancer, non-small cell lung cancer (NSCLC), including inoperable NSCLC or metastatic NSCLC; non-squamous NSCLC, squamous cell lung cancer; small cell lung cancer, including inoperable small cell lung cancer or metastatic small cell lung cancer; early lung cancer before and after definitive treatment; cervical cancer, including metastatic NSCLC. Cervical cancer, early cervical cancer before and after definitive treatment; head and neck tumors, including squamous cell carcinoma of the head and neck; Hodgkin's lymphoma; gastrointestinal tumors, metastatic squamous cell esophageal cancer; bladder cancer, including metastatic urothelial carcinoma and renal cancer; endometrial cancer, including metastatic endometrial cancer and early endometrial cancer before and after definitive treatment; breast cancer, including metastatic breast cancer and early endometrial cancer before and after definitive treatment; liver cancer, including metastatic liver cancer or liver cancer unsuitable for surgery, and early liver cancer before and after definitive treatment; unsuitable solid tumors or metastatic solid tumors, including unsuitable solid tumors or metastatic solid tumors with microsatellite stability.

[0251] In another general aspect, the present invention relates to the use of an aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab for the treatment of malignant tumors, comprising administering a pharmaceutically effective amount of the aqueous pharmaceutical composition described herein.

[0252] In another general aspect, the present invention relates to the use of an aqueous pharmaceutical composition of an anti-PD-1 antibody for the treatment of malignant tumors in vivo in subjects with corresponding needs, the composition comprising:

[0253] (a) Prolgolimab at a concentration of 15 mg / mL to 40 mg / mL as an antibody;

[0254] (b) Trehalose dihydrate at a concentration of 80 mg / mL to 110 mg / mL;

[0255] (c) Sodium acetate trihydrate at a concentration of 0.2 mg / mL to 2.5 mg / mL; and

[0256] (d) Add acetic acid to a pH of 4.5–5.5.

[0257] In some embodiments of the present invention, the concentration of Prolgolimab can be 15 mg / mL to 25 mg / mL.

[0258] In some embodiments of the present invention, the concentration of Prolgolimab can be 20 mg / mL.

[0259] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 95 mg / mL to 105 mg / mL.

[0260] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 100 mg / mL.

[0261] In some embodiments of the present invention, the concentration of sodium acetate trihydrate can be 1.6 mg / mL to 1.9 mg / mL.

[0262] In some embodiments of the present invention, the concentration of sodium acetate trihydrate can be 1.7 mg / mL to 1.8 mg / mL.

[0263] In some embodiments of the present invention, the concentration of sodium acetate trihydrate may be 1.742 mg / mL.

[0264] In some embodiments of the present invention, acetic acid may be added to a pH value of 5.0.

[0265] In some embodiments of the present invention, the concentration of acetic acid may be 0.04 mg / mL to 0.77 mg / mL.

[0266] In some embodiments of the present invention, the concentration of acetic acid may be 0.40 mg / mL to 0.50 mg / mL.

[0267] In some embodiments of the present invention, the concentration of acetic acid may be 0.43 mg / mL.

[0268] In another general aspect, the present invention relates to the use of an aqueous pharmaceutical composition of an anti-PD-1 antibody for the treatment of malignant tumors in vivo in subjects with corresponding needs, the composition comprising:

[0269] (a) Prolgolimab at a concentration of 5 mg / mL to 40 mg / mL as an antibody;

[0270] (b) Trehalose dihydrate at a concentration of 70 mg / mL to 110 mg / mL;

[0271] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0272] (d) L-histidine hydrochloride at a concentration of 0.2–3.5 mg / mL.

[0273] In some embodiments of the present invention, the concentration of Prolgolimab can be 15 mg / mL to 25 mg / mL.

[0274] In some embodiments of the present invention, the concentration of Prolgolimab can be 20 mg / mL.

[0275] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 95 mg / mL to 105 mg / mL.

[0276] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 100 mg / mL.

[0277] In some embodiments of the present invention, the concentration of L-histidine can be 0.7 mg / mL to 1.0 mg / mL.

[0278] In some embodiments of the present invention, the concentration of L-histidine may be 0.92 mg / mL.

[0279] In some embodiments of the present invention, the concentration of L-histidine hydrochloride may be 2.8 mg / mL to 3.3 mg / mL.

[0280] In some embodiments of the present invention, the concentration of L-histidine hydrochloride may be 2.96 mg / mL.

[0281] In some embodiments of the present invention, the pH value of the composition may be 5.5 to 6.5.

[0282] In some embodiments of the present invention, the pH value of the composition may be 5.5.

[0283] In one embodiment, an aqueous pharmaceutical composition providing an anti-PD-1 antibody is used to treat malignant tumors in a subject with corresponding needs, the composition comprising:

[0284] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0285] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0286] (c) Sodium acetate trihydrate at a concentration of 0.2–2.5 mg / mL; and

[0287] (d) Add acetic acid to a pH of 4.5–5.5.

[0288] In one embodiment, an aqueous pharmaceutical composition providing an anti-PD-1 antibody is used to treat malignant tumors in a subject with corresponding needs, the composition comprising:

[0289] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0290] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0291] (c) Sodium acetate trihydrate at a concentration of 1.7 mg / mL to 1.8 mg / mL; and

[0292] (d) Add acetic acid to a pH of 5.0.

[0293] In one embodiment, an aqueous pharmaceutical composition for treating malignant tumors in vivo in a subject with a corresponding need is provided, the composition comprising:

[0294] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0295] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0296] (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL; and

[0297] (d) Add acetic acid to a pH of 5.0.

[0298] In one embodiment, an aqueous pharmaceutical composition providing an anti-PD-1 antibody is used to treat malignant tumors in a subject with corresponding needs, the composition comprising:

[0299] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0300] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0301] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0302] (d) L-histidine hydrochloride at concentrations of 0.2–3.5 mg / mL,

[0303] (e) wherein the pH value of the composition is 5.5 to 6.5.

[0304] In one embodiment, an aqueous pharmaceutical composition providing an anti-PD-1 antibody is used to treat malignant tumors in a subject with corresponding needs, the composition comprising:

[0305] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0306] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0307] (c) L-histidine at a concentration of 0.92 mg / mL; and

[0308] (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL,

[0309] (e) wherein the pH value of the composition is 5.5.

[0310] In one embodiment, an aqueous pharmaceutical composition providing an anti-PD-1 antibody is intended for the treatment of malignant tumors in vivo in a subject with corresponding needs, wherein each 1 mL of the composition contains:

[0311] I:

[0312]

[0313] In one embodiment, an aqueous pharmaceutical composition providing an anti-PD-1 antibody is intended for the treatment of malignant tumors in vivo in a subject with corresponding needs, wherein each 1 mL of the composition contains:

[0314] II:

[0315]

[0316] The aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention may further contain a suitable solubilizer.

[0317] In some embodiments of the present invention, the solubilizer may be poloxamer 188.

[0318] In some embodiments of the present invention, the amount of poloxamer 188 may be greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0319] In some embodiments of the present invention, the amount of poloxamer 188 may be 0 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.4 mg / mL, 0.5 mg / mL, 0.6 mg / mL, 0.7 mg / mL, 0.8 mg / mL, 0.9 mg / mL, or 1.0 mg / mL.

[0320] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administration of the composition at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0321] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administration of the composition at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0322] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administering the composition every two weeks.

[0323] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administration of the composition once every two weeks.

[0324] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administration of the composition every three weeks.

[0325] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administration of the composition once every three weeks.

[0326] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administering the composition every two weeks at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0327] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administration of the composition once every two weeks at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0328] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administration of the composition at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab every three weeks.

[0329] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include administration of the composition once every three weeks at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0330] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include parenteral administration of the composition.

[0331] In some embodiments of the present invention, the parenteral administration may be intravenous, subcutaneous, or intramuscular.

[0332] In some embodiments of the present invention, the use of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab may include intravenous infusion of the composition.

[0333] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention can be administered by intravenous infusion over a period of 60 minutes. In cases of good tolerability, the infusion time can be shortened to 30 minutes.

[0334] In some embodiments of the present invention, the malignant tumor is: melanoma, including inoperable melanoma or metastatic melanoma, and early melanoma before or after definitive treatment; lung cancer, non-small cell lung cancer (NSCLC), including inoperable non-small cell lung cancer or metastatic non-small cell lung cancer.

[0335] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, the composition comprising:

[0336] (a) Prolgolimab at a concentration of 15 mg / mL to 40 mg / mL as an antibody;

[0337] (b) Trehalose dihydrate at a concentration of 80 mg / mL to 110 mg / mL;

[0338] (c) Sodium acetate trihydrate at a concentration of 0.2 mg / mL to 2.5 mg / mL; and

[0339] (d) Add acetic acid to a pH of 4.5–5.5.

[0340] In some embodiments of the present invention, the concentration of Prolgolimab can be 15 mg / mL to 25 mg / mL.

[0341] In some embodiments of the present invention, the concentration of Prolgolimab can be 20 mg / mL.

[0342] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 95 mg / mL to 105 mg / mL.

[0343] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 100 mg / mL.

[0344] In some embodiments of the present invention, the concentration of sodium acetate trihydrate can be 1.6 mg / mL to 1.9 mg / mL.

[0345] In some embodiments of the present invention, the concentration of sodium acetate trihydrate can be 1.7 mg / mL to 1.8 mg / mL.

[0346] In some embodiments of the present invention, the concentration of sodium acetate trihydrate may be 1.742 mg / mL.

[0347] In some embodiments of the present invention, acetic acid may be added to a pH value of 5.0.

[0348] In some embodiments of the present invention, the concentration of acetic acid may be 0.04 mg / mL to 0.77 mg / mL.

[0349] In some embodiments of the present invention, the concentration of acetic acid may be 0.40 mg / mL to 0.50 mg / mL.

[0350] In some embodiments of the present invention, the concentration of acetic acid may be 0.43 mg / mL.

[0351] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, the composition comprising:

[0352] (a) Prolgolimab at a concentration of 5 mg / mL to 40 mg / mL as an antibody;

[0353] (b) Trehalose dihydrate at a concentration of 70 mg / mL to 110 mg / mL;

[0354] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0355] (d) L-histidine hydrochloride at a concentration of 0.2–3.5 mg / mL.

[0356] In some embodiments of the present invention, the concentration of Prolgolimab can be 15 mg / mL to 25 mg / mL.

[0357] In some embodiments of the present invention, the concentration of Prolgolimab can be 20 mg / mL.

[0358] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 95 mg / mL to 105 mg / mL.

[0359] In some embodiments of the present invention, the concentration of trehalose dihydrate can be 100 mg / mL.

[0360] In some embodiments of the present invention, the concentration of L-histidine can be 0.7 mg / mL to 1.0 mg / mL.

[0361] In some embodiments of the present invention, the concentration of L-histidine may be 0.92 mg / mL.

[0362] In some embodiments of the present invention, the concentration of L-histidine hydrochloride may be 2.8 mg / mL to 3.3 mg / mL.

[0363] In some embodiments of the present invention, the concentration of L-histidine hydrochloride may be 2.96 mg / mL.

[0364] In some embodiments of the present invention, the pH value of the composition may be 5.5 to 6.5.

[0365] In some embodiments of the present invention, the pH value of the composition may be 5.5.

[0366] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, the composition comprising:

[0367] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0368] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0369] (c) Sodium acetate trihydrate at a concentration of 0.2–2.5 mg / mL; and

[0370] (d) Add acetic acid to a pH of 4.5–5.5.

[0371] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, the composition comprising:

[0372] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0373] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0374] (c) Sodium acetate trihydrate at a concentration of 1.7 mg / mL to 1.8 mg / mL; and

[0375] (d) Add acetic acid to a pH of 5.0.

[0376] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, the composition comprising:

[0377] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0378] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0379] (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL; and

[0380] (d) Add acetic acid to a pH of 5.0.

[0381] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, the composition comprising:

[0382] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0383] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0384] (c) L-histidine at a concentration of 0.2–2.5 mg / mL; and

[0385] (d) L-histidine hydrochloride at concentrations of 0.2–3.5 mg / mL,

[0386] (e) wherein the pH value of the composition is 5.5 to 6.5.

[0387] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, the composition comprising:

[0388] (a) Prolgolimab at a concentration of 20 mg / mL as an antibody;

[0389] (b) Trehalose dihydrate at a concentration of 100 mg / mL;

[0390] (c) L-histidine at a concentration of 0.92 mg / mL; and

[0391] (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL,

[0392] (e) wherein the pH value of the composition is 5.5.

[0393] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0394] I:

[0395]

[0396] In one embodiment, a method for treating malignant tumors in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0397] II:

[0398]

[0399] The aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention may further contain a suitable solubilizer.

[0400] In some embodiments of the present invention, the solubilizer may be poloxamer 188.

[0401] In some embodiments of the present invention, the amount of poloxamer 188 may be greater than 0 mg / mL and less than or equal to 1 mg / mL.

[0402] In some embodiments of the present invention, the amount of poloxamer 188 may be 0 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.4 mg / mL, 0.5 mg / mL, 0.6 mg / mL, 0.7 mg / mL, 0.8 mg / mL, 0.9 mg / mL, or 1.0 mg / mL.

[0403] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab at a dose of 1 mg / kg body weight.

[0404] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab aqueous pharmaceutical composition at a dose of 3 mg / kg body weight.

[0405] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab every two weeks.

[0406] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab once every two weeks.

[0407] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab every three weeks.

[0408] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab once every three weeks.

[0409] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab at a dose of 1 mg / kg body weight every two weeks.

[0410] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab aqueous pharmaceutical composition at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab every two weeks.

[0411] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab at a dose of 3 mg / kg body weight every three weeks.

[0412] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab aqueous pharmaceutical composition at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab once every three weeks.

[0413] In some embodiments of the present invention, a method for treating malignant tumors in vivo in a subject with corresponding needs may include parenteral administration of a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab.

[0414] In some embodiments of the present invention, the parenteral administration may be intravenous, subcutaneous, or intramuscular.

[0415] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include intravenous infusion of a therapeutically effective amount of the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab.

[0416] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention can be administered by intravenous infusion over a period of 60 minutes. In cases of good tolerability, the infusion time can be shortened to 30 minutes.

[0417] In some embodiments of the present invention, the malignant tumor is: melanoma, including inoperable melanoma or metastatic melanoma, and early melanoma before or after definitive treatment; lung cancer, non-small cell lung cancer (NSCLC), including inoperable non-small cell lung cancer or metastatic non-small cell lung cancer.

[0418] In one embodiment, a method for treating malignant tumors in a subject with a corresponding need is provided, comprising administering a therapeutically effective amount of Prolgolimab.

[0419] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of Prolgolimab at a dose of 1 mg / kg.

[0420] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of Prolgolimab at a dose of 3 mg / kg.

[0421] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective dose of Prolgolimab every two weeks.

[0422] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective dose of Prolgolimab once every two weeks.

[0423] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective dose of Prolgolimab every three weeks.

[0424] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective dose of Prolgolimab once every three weeks.

[0425] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of Prolgolimab at a dose of 1 mg / kg every two weeks.

[0426] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of Prolgolimab at a dose of 1 mg / kg once every two weeks.

[0427] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of Prolgolimab at a dose of 3 mg / kg every three weeks.

[0428] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include administering a therapeutically effective amount of Prolgolimab at a dose of 3 mg / kg once every three weeks.

[0429] In some embodiments of the present invention, a method for treating malignant tumors in vivo in a subject with corresponding needs may include parenteral administration of a therapeutically effective amount of Prolgolimab.

[0430] In some embodiments of the present invention, the parenteral administration may be intravenous, subcutaneous, or intramuscular.

[0431] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include intravenous infusion of a therapeutically effective amount of Prolgolimab.

[0432] In some implementations, Prolgolimab can be administered via intravenous infusion over 60 minutes. In cases of good tolerability, the infusion time can be reduced to 30 minutes.

[0433] In some embodiments of the present invention, the malignant tumor is: melanoma, including inoperable melanoma or metastatic melanoma, and early melanoma before or after definitive treatment; lung cancer, non-small cell lung cancer (NSCLC), including inoperable non-small cell lung cancer or metastatic non-small cell lung cancer.

[0434] In one embodiment of the present invention, a pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity is provided, wherein each 1 mL of the pharmaceutical composition contains:

[0435]

[0436] In one embodiment of the present invention, a pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity is provided, wherein each 1 mL of the pharmaceutical composition contains:

[0437]

[0438] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity can be administered at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0439] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity may be administered at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0440] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for application to a subject to inhibit PD-1 protein activity may be applied every two weeks.

[0441] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for application to a subject to inhibit PD-1 protein activity can be administered once every two weeks.

[0442] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for application to a subject to inhibit PD-1 protein activity can be administered once every three weeks.

[0443] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity may be administered every two weeks at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0444] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity may be administered once every two weeks at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0445] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity may be administered every three weeks at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0446] In some embodiments of the invention, the aqueous pharmaceutical composition suitable for administration to a subject to inhibit PD-1 protein activity may be administered once every three weeks at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

[0447] In some embodiments of the present invention, a method for treating malignant tumors in vivo in a subject with corresponding needs may include parenteral administration of a therapeutically effective amount of Prolgolimab.

[0448] In some embodiments of the present invention, the parenteral administration may be intravenous, subcutaneous, or intramuscular.

[0449] In some embodiments of the present invention, a method for treating malignant tumors in a subject with a corresponding need may include intravenous infusion of a therapeutically effective amount of Prolgolimab.

[0450] In some implementations, Prolgolimab can be administered via intravenous infusion over 60 minutes. In cases of good tolerability, the infusion time can be reduced to 30 minutes.

[0451] In some embodiments of the present invention, the malignant tumor is: melanoma, including inoperable melanoma or metastatic melanoma, and early melanoma before or after definitive treatment; lung cancer, non-small cell lung cancer (NSCLC), including inoperable non-small cell lung cancer or metastatic non-small cell lung cancer. Attached Figure Description

[0452] The invention will be more readily understood through the following detailed description of embodiments with reference to the accompanying drawings, in which:

[0453] Figure 1 and Figure 2 The figure shows the dynamic changes in the concentration of BCD-100 in the patient's serum (μg / mL) during 6 administrations (adjusted for coefficients) (BCD-100-1 test).

[0454] Figure 3 This is the experimental design diagram for BCD-100-2 / MIRACULUM.

[0455] Figure 4 This is a schematic diagram of the experiment.

[0456] Figure 5 The figure shows the overall survival of patients in the first group (BCD-100, 1 mg / kg, every two weeks) based on the results of the BCD-100-2 / MIRACULUM trial.

[0457] Figure 6 The figure shows the overall survival of patients in the second group (BCD-100, 3 mg / kg, every three weeks) based on the results of the BCD-100-2 / MIRACULUM trial.

[0458] Figure 7 The figure shows progression-free survival in the BCD-100 1 mg / kg group based on the results of the BCD-100-2 / MIRACULUM trial (according to irRECIST guidelines).

[0459] Figure 8The figure shows the progression-free survival of patients in the BCD-100 3 mg / kg group based on the results of the BCD-100-2 / MIRACULUM trial (according to irRECIST guidelines).

[0460] Figure 9 BCD-100 concentration graph for patients who received 1 mg / kg of the product every two weeks after a single dose of BCD-100 (Results of the BCD-100-2 / MIRACULUM Trial).

[0461] Figure 10 BCD-100 concentration graph for patients receiving 3 mg / kg BCD-100 every three weeks (Results of BCD-100-2 / MIRACULUM Trial).

[0462] Figure 11 The figure shows the Th9 portion of the total helper T cell population in a patient group receiving BCD-100 at a dose of 1 mg / kg every two weeks. This figure illustrates the different types of responses to this treatment (BCD-100-2 / MIRACULUM trial results).

[0463] Figure 12 The figure shows the Th9 portion of the total helper T cell population in a patient group that received a 3 mg / kg dose of BCD-100 every three weeks. This figure illustrates the different types of responses to this treatment (results from the BCD-100-2 / MIRACULUM trial). Detailed Implementation

[0464] definition:

[0465] The terms used in this specification generally have their ordinary meaning in the art, corresponding to the context of this invention and the specific context in which each term is used. To provide additional guidance to those skilled in the art, certain terms used in this invention will be described below or in other parts of this specification, and synonyms for certain terms will be given. The listing of one or more synonyms does not preclude the availability of other synonyms. Examples used in any part of this specification, including any terms given herein, are for illustrative purposes only and do not constitute any limitation on the scope or meaning of the invention or any of the exemplified terms. This invention is not limited to the various embodiments given in this specification.

[0466] Unless otherwise stated in this application, the term "monoclonal antibody" as used herein refers to humanized antibodies or fully human antibodies. The monoclonal antibodies of this invention can be obtained, for example, by recombinant technology, phage display technology, synthetic technology, or a combination of these technologies or other technologies well known in the art.

[0467] As used herein, the term “monoclonal antibody cluster” refers to a homogeneous or substantially homogeneous antibody cluster (i.e., antibodies that compete with each other for the same antigen / epitope in enzyme-linked immunosorbent assay (ELISA), or more preferably, antibodies that are identical in terms of amino acid sequence constitute at least 96% or 96% of the cluster, more preferably not less than about 97% or 98%, and even more preferably at least 99%).

[0468] Native full-length antibodies are immunoglobulin molecules consisting of four polypeptide chains linked by disulfide bonds: two heavy (H) chains, approximately 50–70 kDa in full length, and two light (L) chains, approximately 25 kDa in full length. The amino-terminal portion of each chain contains a variable region consisting of approximately 100–110 or more amino acids for binding the antigen. The carboxyl-terminal region of each chain forms a constant region primarily responsible for effector function. Light chains are classified into κ and λ types and possess specific constant regions. Each light chain is characterized by an N-terminal light chain variable region (hereinafter referred to as VL or VK) and a light chain constant region (CL or CK) consisting of a single region. Heavy chains are classified into five types: γ, δ, α, μ, and ε, and each defines one of five immunoglobulin classes: IgG, IgM, IgA, IgD, and IgE. Some immunoglobulin types can be further subdivided into subtypes (isotypes) such as IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. Each heavy chain type is characterized by its specific constant region Fc. Each heavy chain includes an N-terminal variable region (hereinafter referred to as VH) and a constant region CH. The heavy chain constant regions of IgG, IgD, and IgA consist of three regions (CH1, CH2, and СН3), while the heavy chain constant regions of IgM and IgE consist of four regions (CH1, CH2, CH3, and СН4). VH and VL can also be divided into so-called hypervariable regions (complementarity-determining regions, CDRs), which are distributed among the more conserved backbone regions (FRs). Each variable region includes three CDRs and four FRs, in the following order from N-terminus to C-terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4.

[0469] Each pair of variable regions on the light and heavy chains forms an antigen-binding site for the antibody. Therefore, a complete IgG antibody has two binding sites. These two binding sites are identical except for bifunctional or bispecific antibodies. In this document, the terms "antigen-binding moiety," "antigen-binding region," and "antigen-binding domain" are used interchangeably when referring to an antibody region containing amino acid residues that interact with the antigen and confer specificity and affinity to the antigen. This antibody fragment contains the backbone amino acid residues necessary for the antigen-binding residues to maintain their correct conformation.

[0470] "Antibody fragment" can be an antibody fragment or an antibody fragment with full-length antibody activity. Antibody fragments can be F(ab')2, F(ab)2, Fab', Fab Fv, and scFv.

[0471] In this application, the terms "inhibition" or "suppression" used when referring to the antibody activity of the present invention refer to the ability to significantly block, prevent, limit, slow down, stop, reduce, or reverse the development or severity of the inhibited target, including but not limited to biological activities (e.g., PD-1 activity) or characteristics, diseases, or conditions. The degree of inhibition or suppression of PD-1 activity resulting from the binding of the antibody of the present invention to PD-1 is preferably at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or higher.

[0472] When referring to nucleic acid or protein products (such as antibodies), the terms "separated" or "isolated" mean that the nucleic acid or protein molecule has been isolated from at least one contaminant that is normally associated with it in its natural source and has been identified. Preferably, "isolated antibody" refers to an antibody that is substantially free of other antibodies with specific antigen specificity (e.g., the pharmaceutical compositions of the present invention contain an isolated antibody that specifically binds to PD-1, but substantially free of antibodies that specifically bind to antigens other than PD-1).

[0473] The functional linking of polynucleotides with other polynucleotides is called "functional binding". For example, a promoter or enhancer can functionally bind to the coding sequence it is involved in transcribing. If the coding polynucleotides of two polypeptides (preferably located within the same open reading frame) are functionally bound, then one polypeptide can "functionally bind" to the other polypeptide.

[0474] The term "specific binding" between an antibody and its antigen target (antigen) refers to immune specificity. If an antibody binds more strongly to a particular antigenic epitope than to other antigenic epitopes, then the antibody can be said to specifically bind to that antigenic target. Specific binding does not preclude cross-reactivity with other antigens that have similar antigenic epitopes.

[0475] The VL region in the antibody of the present invention can be either λ-type VL or κ-type VL. The term "VL region" simultaneously covers both λ and κ isotypes of VL containing one or more amino acid substitutions, insertions, or deletions.

[0476] The term "pharmaceutical composition" refers to a composition and / or formulation containing a therapeutically effective amount of the antibody of the present invention and excipients (carriers, diluents, mediators, solvents and other excipients).

[0477] As used in this application, the term "buffer solution" refers to an aqueous solution containing a mixture of an acid (usually a weak acid, such as acetic acid or citric acid) and its conjugate base (such as acetates or citrates like sodium acetate or sodium citrate, and hydrates of such salts like sodium acetate trihydrate), or an aqueous solution containing a base (usually a weak base, such as histidine) and its conjugate acid (such as histidine hydrochloride). Due to the "buffering effect" imparted by the "buffer," the pH of the "buffer solution" changes only slightly when a small amount of strong base or strong acid is added, or when it is diluted or concentrated.

[0478] In this document, a “buffer system” comprises one or more buffers and / or their acid / base conjugates, more preferably one or more buffers and their acid / base conjugates, and most preferably one buffer and its acid / base conjugate. Unless otherwise stated, any concentration (buffer concentration) mentioned herein when referring to a “buffer system” may appropriately refer to the total concentration of the buffer and / or its acid / base conjugate. That is, the concentration mentioned herein when referring to a “buffer system” may refer to the total concentration of all relevant buffering substances (i.e., substances that have reached dynamic equilibrium with each other, such as citrate / citric acid). The total pH of the composition containing the relevant buffer system reflects the equilibrium concentration of all relevant buffering substances (i.e., the equilibrium between the buffer and its acid / base conjugate).

[0479] The term "buffer" as used herein refers to the acid-base component (typically a weak acid or weak base) in a buffer solution or buffer buffer. Buffers help maintain the pH of a given solution at or near a predetermined value, and are often used to adjust the pH to a predetermined value. A buffer can be a single compound capable of producing the desired buffering effect, especially when mixed with an appropriate amount of a corresponding "acid / base conjugate" (and having the appropriate ability to undergo proton exchange with it).

[0480] As used herein, the term "solvent" refers to a pharmaceutically acceptable nonionic surfactant. Solubilizers may be used alone or in combination. Examples of solubilizers include, but are not limited to, polysorbate 20 or polysorbate 80, poloxamer 184 or poloxamer 188, or pranoxamer.

[0481] As used herein, the terms "permeabilizer," "toning agent," and "osmotic regulator" refer to excipients that enable a liquid antibody solution to achieve the desired osmotic pressure. In some embodiments, a toning agent can increase the osmotic pressure of a liquid antibody formulation to isotonicity, thereby making the liquid antibody formulation compatible with the cellular physiology of the target biological tissue. In another embodiment, a toning agent can help improve antibody stability. An "isotonic" drug is a drug with an osmotic pressure equal to that of human blood. The osmotic pressure of an isotonic drug is typically about 250–350 mOsm / kg. The term "hypotonic" is used to describe formulations with an osmotic pressure lower than that of human blood. Correspondingly, the term "hypertonic" is used to describe formulations with an osmotic pressure higher than that of human blood. Isotonicity can be measured, for example, by a vapor osmoremeter or a freezing point osmoremeter. The penetrant can be: enantiomer (e.g., L-enantiomer or D-enantiomer) or racemic form; α or β isomer, including α / α, β / β, α / β or β / α; free acid or free base form; salt form; hydrated form (e.g., monohydrate); or anhydrous form. Penetrants are, for example, but not limited to, sugars (trehalose dihydrate, sucrose, glucose), polyols (mannitol, sorbitol), amino acids (proline, arginine, glycine), or salts (sodium chloride, potassium chloride, magnesium chloride).

[0482] The terms "long-term storage" and "long-term stability" are intended to indicate that the corresponding pharmaceutical composition can be stored for three months or longer, six months or longer, preferably one year or longer, and most preferably with a stable shelf life of at least two years. Generally, "long-term storage" and "long-term stability" also imply that the length of stable storage time during which the formulation is rendered unusable for its target pharmaceutical purpose without degradation of stability is at least equivalent to or better than the stable shelf life typically required for currently commercially available formulations of the anti-PD-1 antibody Prolgolimab.

[0483] The term “parenteral administration” refers to administration regimens that are usually performed by injection, and in particular include intravenous, intramuscular, intra-arterial, intratracheal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, tracheal, subcutaneous, intra-articular, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injections or infusions.

[0484] The term "use" refers to the ability of the antibody of the present invention or a pharmaceutical composition containing the antibody of the present invention to treat, slow down, accelerate remission, or reduce the recurrence rate of a disease or condition mediated by a receptor that the antibody of the present invention can bind. Examples of diseases include, but are not limited to, malignant tumors, including: melanoma, including inoperable melanoma or metastatic melanoma, and early melanoma before and after definitive treatment; lung cancer, non-small cell lung cancer (NSCLC), including inoperable NSCLC or metastatic NSCLC; non-squamous NSCLC, squamous cell lung cancer; small cell lung cancer, including inoperable small cell lung cancer or metastatic small cell lung cancer; early lung cancer before and after definitive treatment; cervical cancer, including metastatic cervical cancer, and early cervical cancer before and after definitive treatment; head and neck tumors. This includes squamous cell carcinoma of the head and neck; Hodgkin's lymphoma; gastrointestinal tumors, including metastatic squamous cell esophageal cancer; bladder cancer, including metastatic urothelial carcinoma and renal cancer; endometrial cancer, including metastatic endometrial cancer and early endometrial cancer before and after definitive treatment; breast cancer, including metastatic breast cancer and early endometrial cancer before and after definitive treatment; liver cancer, including metastatic liver cancer or inoperable liver cancer, and early liver cancer before and after definitive treatment; and inoperable solid tumors or metastatic solid tumors, including inoperable solid tumors or metastatic solid tumors with microsatellite stability.

[0485] The term "treatment method" refers to the ability of the antibody of the present invention or a pharmaceutical composition containing the antibody of the present invention to treat a disease or condition associated with PD-1 activity, slow its progression, accelerate its remission, or reduce its relapse rate. "Treatment" of a disease, condition, or symptom may include: preventing or delaying the onset of clinical symptoms of a disease, condition, or symptom developing in the body; suppressing a disease, condition, or symptom, i.e., stopping, reducing, or delaying its development, relapse (in the case of maintenance therapy), or at least one clinical or subclinical symptom; or alleviating or relieving a disease, i.e., causing the disease, condition, or symptom to subside. Examples of diseases include, but are not limited to, malignant tumors, including: melanoma, including inoperable melanoma or metastatic melanoma, early melanoma before or after definitive surgical treatment; lung cancer, non-small cell lung cancer (NSCLC), including inoperable non-small cell lung cancer or metastatic non-small cell lung cancer.

[0486] As used herein, the term "aqueous composition" refers to a water-based composition in which the water may be: water; water for injection; physiological saline (a 0.9% to 1.0% aqueous solution of sodium chloride).

[0487] In one embodiment of the invention, the treatment subject, also referred to as the patient, is a mammal, preferably a human. The subject can also be a male or female of any age.

[0488] In this specification, the words “including,” “having,” “comprising,” or their variations such as “having,” “having,” “containing,” and all grammatical variations shall be understood to mean that one or a group of complete objects are included in the description, but do not exclude any other one or a group of complete objects.

[0489] Example of implementation :

[0490] This invention relates to suitable aqueous pharmaceutical compositions of the anti-PD-1 antibody Prolgolimab. In one embodiment of the invention, an aqueous pharmaceutical composition of Prolgolimab may contain an acetate-based buffer and trehalose. Poloxamer 188 may be added as a solubilizer. In another embodiment of the invention, an aqueous pharmaceutical composition of Prolgolimab may contain a histidine-based buffer and trehalose. Poloxamer 188 may be added as a solubilizer.

[0491] The histidine-based buffer can be obtained by combining L-histidine with histidine hydrochloride, or further with hydrochloric acid or other acids. It is understood that while histidine hydrochloride can be used as a salt of the histidine-based buffer, any other histidine-based salt may also be used in the histidine-based buffer without departing from the scope of this invention.

[0492] The acetate-based buffer can be obtained by combining acetic acid with sodium acetate trihydrate. It is understood that while sodium acetate trihydrate can be used as the salt of the acetate-based buffer, any other acetate, such as potassium acetate, can also be used in the acetate-based buffer without departing from the scope of this invention.

[0493] In addition, the compositions of the present invention may contain one or more other suitable excipients known to those skilled in the art.

[0494] In some embodiments, the liquid pharmaceutical composition is stored stably in the sense that it does not undergo further protein aggregation or modification compared to the stability index at time zero.

[0495] In one embodiment, surprisingly, the inventors have obtained a high-concentration aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab, wherein the concentration of Prolgolimab can be from 90 mg / mL to 150 mg / mL. In some embodiments of the present invention, the concentration of Prolgolimab can be 90 mg / mL, 95 mg / mL, 100 mg / mL, 105 mg / mL, 110 mg / mL, 115 mg / mL, 120 mg / mL, 125 mg / mL, 130 mg / mL, 135 mg / mL, 140 mg / mL, 145 mg / mL, or 150 mg / mL.

[0496] The high-concentration aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of this invention exhibits colloidal stability under vigorous stirring (800 rpm) for 120 hours, high thermal stability under heating conditions of 50°C and 37°C, and a viscosity of less than 50 cP suitable for parenteral administration.

[0497] The above composition is suitable for parenteral administration, such as intravenous, subcutaneous, intradermal, intra-arterial, intrathecal, intraperitoneal, intra-articular, and / or intramuscular administration.

[0498] The pharmaceutical compositions of the present invention can be administered to individuals in need of treatment by: systemic injection, such as intravenous, subcutaneous or intramuscular injection; injection or application to a suitable site, such as direct injection or application to the site when surgery is possible; or local application.

[0499] The above composition can be administered to individuals requiring intravenous infusion.

[0500] In some embodiments, the aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab of the present invention can be administered by intravenous infusion over a period of 60 minutes. In cases of good tolerability, the infusion time can be shortened to 30 minutes.

[0501] The aqueous pharmaceutical composition of the anti-PD-1 antibody Prolgolimab can be used after dilution. The desired volume of the composition is transferred from a vial to an infusion container containing a sterile 0.9% sodium chloride solution or a sterile 5% dextran solution. The concentration of the composition in the resulting solution can be from 0.5 mg / mL to 10 mg / mL. To avoid foaming, the resulting solution is stirred by gently inverting the infusion container.

[0502] Treatment methods and uses of aqueous compositions

[0503] In another embodiment, the present invention relates to a treatment method for mammals, comprising administering to the mammal a therapeutically effective amount of the pharmaceutical composition of the present invention, wherein the concentration of the anti-PD-1 antibody Prolgolimab is 15 mg / mL to 40 mg / mL, and the mammal may suffer from a disease or condition that can be effectively treated with the anti-PD-1 antibody Prolgolimab of the present invention.

[0504] In another embodiment, the present invention relates to a treatment method for mammals, comprising administering to the mammal a therapeutically effective amount of the pharmaceutical composition of the present invention, wherein the concentration of the anti-PD-1 antibody Prolgolimab is 20 mg / mL, and the mammal may suffer from a disease or condition that can be effectively treated with the anti-PD-1 antibody Prolgolimab of the present invention.

[0505] In a preferred embodiment, the mammal is a human.

[0506] In another embodiment, the present invention relates to a method of treating a subject with a corresponding need, comprising administering to the subject a therapeutically effective amount of the pharmaceutical composition of the present invention, wherein the concentration of the anti-PD-1 antibody Prolgolimab is 15 mg / mL to 40 mg / mL, and the subject may suffer from a disease or condition that can be effectively treated with the anti-PD-1 antibody Prolgolimab of the present invention.

[0507] In another embodiment, the present invention relates to a method of treating a subject with a corresponding need, comprising administering to the subject a therapeutically effective amount of the pharmaceutical composition of the present invention, wherein the concentration of the anti-PD-1 antibody Prolgolimab is 20 mg / mL, and the subject may suffer from a disease or condition that can be effectively treated with the anti-PD-1 antibody Prolgolimab of the present invention.

[0508] In a preferred embodiment, the object is a person.

[0509] In one such embodiment, the present invention relates to a melanoma treatment method comprising administering one of the compositions of the invention to a subject in need at a therapeutically effective amount, wherein the concentration of the anti-PD-1 antibody Prolgolimab of the present invention is 15 mg / mL to 40 mg / mL.

[0510] In one such embodiment, the present invention relates to a melanoma treatment method comprising administering one of the compositions of the invention to a subject in need at a therapeutically effective amount, wherein the concentration of the anti-PD-1 antibody Prolgolimab of the present invention is 20 mg / mL.

[0511] In one such embodiment, the present invention relates to a treatment method for inoperable melanoma, comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 15 mg / mL to 40 mg / mL.

[0512] In one such embodiment, the present invention relates to a treatment method for inoperable melanoma, comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 20 mg / mL.

[0513] In one such embodiment, the present invention relates to a treatment method for metastatic melanoma, comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 15 mg / mL to 40 mg / mL.

[0514] In one such embodiment, the present invention relates to a treatment method for metastatic melanoma, comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 20 mg / mL.

[0515] In one such embodiment, the present invention relates to a method for treating early melanoma before and after definitive treatment, comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject with corresponding need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 15 mg / mL to 40 mg / mL.

[0516] In one such embodiment, the present invention relates to a method for treating early melanoma before and after definitive treatment, comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject with a corresponding need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 20 mg / mL.

[0517] In one such embodiment, the present invention relates to a lung cancer treatment method comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 15 mg / mL to 40 mg / mL.

[0518] In one such embodiment, the present invention relates to a lung cancer treatment method comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 20 mg / mL.

[0519] In one such embodiment, the present invention relates to a treatment method for non-small cell lung cancer (NSCLC) comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 15 mg / mL to 40 mg / mL.

[0520] In one such embodiment, the present invention relates to a treatment method for non-small cell lung cancer (NSCLC) comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 20 mg / mL.

[0521] In one such embodiment, the present invention relates to a method for treating non-small cell lung cancer that is unsuitable for surgery or metastatic non-small cell lung cancer, comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject with corresponding need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 15 mg / mL to 40 mg / mL.

[0522] In one such embodiment, the present invention relates to a method for treating non-small cell lung cancer that is unsuitable for surgery or metastatic non-small cell lung cancer, comprising administering one of the present invention compositions of the anti-PD-1 antibody Prolgolimab to a subject in need at a therapeutically effective amount, wherein the concentration of the present invention's anti-PD-1 antibody Prolgolimab is 20 mg / mL.

[0523] In one embodiment, a method for treating melanoma in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0524] I:

[0525]

[0526] In one embodiment, a method for treating melanoma in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0527] II:

[0528]

[0529] In one embodiment, a method for treating inoperable melanoma in a patient with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0530] I:

[0531]

[0532] In one embodiment, a method for treating inoperable melanoma in a patient with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0533] II:

[0534]

[0535]

[0536] In one embodiment, a method for treating metastatic melanoma in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0537] I:

[0538]

[0539] In one embodiment, a method for treating metastatic melanoma in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0540] II:

[0541]

[0542] In one embodiment, a method for treating early-stage melanoma in a subject before and after definitive treatment is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0543] I:

[0544]

[0545] In one embodiment, a method for treating early-stage melanoma in a subject before and after definitive treatment is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0546] II:

[0547]

[0548] In one embodiment, a method for treating lung cancer in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0549] I:

[0550]

[0551] In one embodiment, a method for treating lung cancer in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0552] II:

[0553]

[0554] In one embodiment, a method for treating non-small cell lung cancer (NSCLC) in vivo in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0555] I:

[0556]

[0557] In one embodiment, a method for treating non-small cell lung cancer (NSCLC) in vivo in a subject with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0558] II:

[0559]

[0560] In one embodiment, a method for treating inoperable or metastatic non-small cell lung cancer in a patient with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0561] I:

[0562]

[0563]

[0564] In one embodiment, a method for treating inoperable or metastatic non-small cell lung cancer in a patient with corresponding needs is provided, comprising administering a therapeutically effective amount of an aqueous pharmaceutical composition of an anti-PD-1 antibody, wherein each 1 mL of the composition contains:

[0565] II:

[0566]

[0567] In one such embodiment, the present invention relates to a melanoma treatment method comprising administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab to a subject in need.

[0568] In one such embodiment, the present invention relates to a treatment method for inoperable melanoma, comprising administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab to a subject with appropriate need.

[0569] In one such embodiment, the present invention relates to a treatment method for metastatic melanoma, comprising administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab to a subject in need.

[0570] In one such embodiment, the present invention relates to a method for treating early melanoma before and after definitive treatment, comprising administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab to a subject with the corresponding need.

[0571] In one such embodiment, the present invention relates to a lung cancer treatment method comprising administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab to a subject in need.

[0572] In one such embodiment, the present invention relates to a treatment method for non-small cell lung cancer (NSCLC) comprising administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab to a subject in need.

[0573] In one such embodiment, the present invention relates to a method for treating non-small cell lung cancer that is not suitable for surgery or metastatic non-small cell lung cancer, comprising administering a therapeutically effective amount of the anti-PD-1 antibody Prolgolimab to a subject in need.

[0574] The therapeutically effective dose of the anti-PD-1 antibody and the aqueous composition containing the anti-PD-1 antibody Prolgolimab of this invention depends on the specific condition being treated, the severity of the condition, previous treatments received, and the patient's medical history and response to the treatment. The attending physician may decide to adjust the appropriate dosage so that it can be administered to the patient via a single or multiple injections.

[0575] In one such embodiment, the effective amount of each dose of anti-PD-1 antibody for a patient is about 0.01 to 10 mg / kg body weight, or about 1 to 10 mg / kg body weight, or about 0.05 mg / kg body weight, or about 0.25 mg / kg body weight, or about 0.5 mg / kg body weight, or about 1 mg / kg body weight, or about 2 mg / kg body weight, or about 3 mg / kg body weight, or about 4 mg / kg body weight, or about 5 mg / kg body weight, or about 6 mg / kg body weight, or about 7 mg / kg body weight, or about 8 mg / kg body weight, or about 9 mg / kg body weight, or about 10 mg / kg body weight.

[0576] The frequency of administration can usually be about once a week, or about once every two weeks, or about once every three weeks.

[0577] In another embodiment, the acceptable dose for infusion administration may be: 5–450 mg / dose, or 40 mg, 50 mg, or 60 mg / dose; or 70 mg, 80 mg, 90 mg, or 100 mg / dose; or 110 mg, 120 mg, 130 mg, or 140 mg / dose; or 150 mg, 160 mg, 170 mg, or 180 mg / dose; or 190 mg, 200 mg, 210 mg, or 220 mg / dose. mg / dose; or 230mg, 240mg, 250mg or 260mg / dose; or 270mg, 280mg or 290mg / dose; or 300mg, 310mg, 320mg, 330mg, 340mg or 350mg / dose; or 360mg, 370mg, 380mg, 390mg, 400mg, 410mg, 420mg, 430mg, 440mg or 450mg / dose.

[0578] In one embodiment of the invention, a dose may be delivered by one or more infusions. A dose may be delivered by one, two, or three infusions. In some embodiments of the invention, one to multiple infusions may be administered during treatment. In some embodiments of the invention, the patient's condition may be improved by extending the treatment duration. In some embodiments of the invention, the treatment duration may continue until disease progression or for life.

[0579] In another embodiment, the pharmaceutical composition of the present invention can be prepared in bulk form, wherein the content of each component of the pharmaceutical composition is substantially higher than the amount required for application. Therefore, appropriate dilution is required before application.

[0580] Alternatively, the pharmaceutical composition may be frozen, spray-dried, or lyophilized and then rehydrated in a suitable sterile container before application. Lyophilization can be accomplished using techniques already available in the art that include various steps such as freezing, annealing, primary drying, and secondary drying.

[0581] The pharmaceutical composition can be administered as a single therapeutic agent or in combination with other therapeutic agents as needed. Accordingly, in one embodiment, the above-described treatment and / or prevention methods can be used in conjunction with the administration of a therapeutically effective amount of another active agent. This other active agent can be administered before, after, or during the administration of the pharmaceutical composition disclosed herein. Furthermore, this other active agent can be administered as part of the above-described composition or as a standalone formulation.

[0582] The pharmaceutical composition of the present invention can be administered via various routes, including parenteral, oral, buccal, nasal, rectal, and topical administration. Parenteral administration may include, but is not limited to, transdermal, subcutaneous, intravenous, intra-arterial, intraperitoneal, intradermal, intracardiac, intraventricular, intracranial, intratracheal, intrathecal, intramuscular, intramuscular, and intravitreal injections.

[0583] The pharmaceutical compositions of this invention are particularly suitable for parenteral administration, i.e., subcutaneous, intramuscular, intravenous, intraperitoneal, intraspinal, intra-articular, intrasynovial, and / or intrathecal administration. Parenteral administration can be achieved by bolus injection or continuous infusion. When used for injection, the pharmaceutical compositions can be in standard dosage forms, such as, but not limited to, ampoules, vials, pre-filled syringes, or multi-dose containers with added preservatives. Furthermore, various newly developed drug delivery methods, such as BD Physioject, are also suitable. TM , Injection pens and and The pharmaceutical compositions of the present invention are also suitable for administration via needle-free injection devices and other novel methods. Furthermore, the pharmaceutical compositions of the present invention are suitable for various routes of administration not currently available.

[0584] See also Langer, 1990, Science, 249: 1527-1533.

[0585] The pharmaceutical compositions of the present invention can also be formulated into long-acting formulations. Such long-acting formulations can be administered by implantation (e.g., subcutaneous or intramuscular) or by intramuscular injection. Accordingly, the compositions can be modified, for example, by suitable polymers or hydrophobic materials (e.g., as emulsions in acceptable oil formulations) or ion exchange resins, or modified into slightly soluble derivatives such as slightly soluble salts.

[0586] Such pharmaceutical compositions can be placed in vials, packages, or dispensing devices as needed, which may contain one or more unit dosage forms containing the aforementioned active ingredient. In one embodiment, the dispensing device may include a syringe containing a single, readily injectable dose of the liquid composition. Furthermore, the syringe may be accompanied by instructions for use.

[0587] In another embodiment, the present invention relates to a kit or container containing the aqueous pharmaceutical composition of the present invention. The antibody concentration in the aqueous pharmaceutical composition may vary over a wide range, but generally falls within the range of about 1 mg / mL to about 200 mg / mL. Similarly, the kit may be accompanied by instructions for use.

[0588] The method for obtaining the above composition includes: adding an acetate buffer to an aqueous phase; and then adding the following components in any order: trehalose; Prolgolimab; and / or a solubilizer selected from the group consisting of polysorbate 20, polysorbate 80, poloxamer 188, or a combination thereof.

[0589] The method for obtaining the above composition includes: adding a histidine buffer to an aqueous phase; and then adding the following components in any order: trehalose; Prolgolimab; and / or a solubilizer selected from the group consisting of polysorbate 20, polysorbate 80, poloxamer 188, or a combination thereof.

[0590] Example study:

[0591] The following describes illustrative studies of the reagents and their concentrations used to determine the aqueous pharmaceutical composition for preparing the anti-PD-1 antibody Prolgolimab of the present invention.

[0592] The following examples and studies are for illustrative purposes and are intended to demonstrate the suitability of certain components used in the aqueous pharmaceutical compositions of the anti-PD-1 antibody Prolgolimab. It is understood that other methods and techniques may be used by those skilled in the art without departing from the spirit of the invention.

[0593] The suitability of the aqueous compositions of the present invention was tested by the illustrative methods described below.

[0594] Example 1: Preparation of a highly stable anti-PD-1 antibody formulation, Prolgolimab

[0595] Antibody samples (5 mg / mL) were prepared under pressure using a Stirred Cell (Millipore) apparatus. Initially, the initial antibody formulation was placed in the tank, and then the protein was concentrated to 10 mg / mL using a compressed air stream under continuous stirring. Subsequently, at least 10 volumes of the target formulation aqueous solution containing buffer and permeabilizer, and further containing water-soluble stabilizers if necessary, were added to the tank. After permeation, the antibody was concentrated to approximately 10 mg / mL and removed from the tank for precise protein concentration measurement using UV spectroscopy. Then, the appropriate excipient solution was added to the sample to prepare a solution with a target protein concentration of 5 ± 0.2 mg / mL.

[0596] Protein samples at concentrations of 50 mg / mL or higher are prepared using a Pellicon filter cartridge (Millipore) in tangential flow mode. The initial antibody formulation is first placed in the percolation tank. After the protein is concentrated to 50–100 mg / mL, at least 10 volumes of the target formulation solution containing buffer and permeabilizer, and further containing water-soluble stabilizers if necessary, are added to the system. Alternatively, a concentrate of permeabilizer and water-soluble stabilizers can be added after percolation. After percolation, the antibody is concentrated to a concentration higher than the target concentration and removed from the system to measure the precise protein concentration. Subsequently, a solution with the target protein concentration is prepared by adding the appropriate excipient solution to the sample.

[0597] When a solubilizer is required in the formulation, after percolation and concentration, a surfactant concentrate is added first, and then the antibody is finally diluted to the target concentration by adding an excipient solution.

[0598] The antibody solution is also filtered using a 0.22 μm filter membrane before being finally filled into a sterile container (such as a glass / plastic tube, vial, or syringe).

[0599] Example 2: Determination of protein concentration in test samples

[0600] Protein concentration was measured by ultraviolet spectroscopy at a wavelength of 280 nm using a UV transparent plate.

[0601] First, each sample was diluted to approximately 0.5 mg / mL with the corresponding excipient solution, and then 150 μL of the diluted sample was added to the wells of a UV spectroscopy plate. Subsequently, the absorbance of the solution in each well was measured at a wavelength of 280 nm using a spectrophotometer. The corresponding excipient solution was used as a reference solution.

[0602] Protein (C) concentration (mg / mL) is calculated using the following formula:

[0603] in,

[0604] A 280 The absorbance value at a wavelength of 280 nm;

[0605] ε is the extinction coefficient of the tested protein;

[0606] b is the total dilution factor of the sample;

[0607] l is the thickness of the liquid layer inside the plate well. At 150 μL, l = 0.42 cm.

[0608] Example 3: PEG Aggregation Study

[0609] A test excipient composition solution with a PEG6000 mass concentration of 20-25% was prepared, and the resulting solution was filtered through a Durapore 0.45μm filter.

[0610] The calculated sample, excipient solution, and 20–25% PEG6000 solution were transferred into a 96-well UV plate to ensure that the PEG6000 concentration in some wells was 0%–18% and the protein concentration in each well was 1 mg / mL. All solutions in the wells were thoroughly mixed using pipetting.

[0611] After visually assessing the turbidity of the solution, the absorbance of the solution at a wavelength of 400 nm was measured.

[0612] The presence of PEG leads to a volume substitution effect, where the polymer displaces the protein from its occupied solvent space. This effect causes the protein concentration to gradually increase, and precipitation occurs when the protein concentration exceeds its solubility. The more unstable the sample, the lower the PEG6000 concentration required for the formation of visible aggregates (emulsions).

[0613] Example 4: Evaluation of Colloid Stability by Shaking Test

[0614] The test sample was evenly divided into two 200 μL portions and placed in small glass vials. One vial of each formulation was stored in a refrigerator at 2–8°C, while the other vial was shaken at 800 rpm for a specified time at 2–8°C. After this, each vial was vortexed and then transferred for analysis.

[0615] Example 5: Evaluation of colloid stability using low-temperature concentration method

[0616] The test samples were divided into two portions and placed in small plastic vials. One vial of each formulation was stored in a refrigerator at 2–8°C, and the other in a freezer at -16–20°C for a specified period. Subsequently, the vials were removed from the freezer and allowed to stand at room temperature until the contents were completely thawed. The resulting solutions were then vortexed and analyzed.

[0617] Example 6: Evaluation of thermal stability using the thermal stress method

[0618] The test samples were divided into two parts and placed in separate glass vials. One vial of each composition was stored in a refrigerator at 2–8°C, while the other was incubated in a thermostat at the desired temperature for a specified time. After heating, the vials were removed from the thermostat and allowed to stand at room temperature for approximately 15 minutes before being transferred for analysis.

[0619] Example 7: Determination of Sample Homogeneity by Size Exclusion High Performance Liquid Chromatography (SEC HPLC)

[0620] Tosoh TSK-GelG3000SW XL Chromatographic column, 7.8 mm (inner diameter) × 30 cm, catalog number 08541;

[0621] Column temperature: 25℃;

[0622] Mobile phase rate: 0.7 mL / min;

[0623] Injection volume: 10 μL;

[0624] Sample concentration: 5 mg / mL;

[0625] Detection wavelength: 220nm;

[0626] Wash-off time: 25 minutes;

[0627] Mobile phase: Anhydrous disodium hydrogen phosphate 7.1 mg / mL

[0628] Sodium chloride 17.54 mg / mL.

[0629] The pH of the mobile phase was adjusted to 7.0 using orthophosphoric acid.

[0630] The change in purity after treatment is calculated using the following formula:

[0631] Δ = (Percentage of main peak after treatment – ​​Percentage of main peak before treatment)

[0632] Example 8: Measurement of charge heterogeneity (charge distribution characteristics) of a sample using Labchip GXII (Caliper)

[0633] Preparation of test samples

[0634] After diluting the sample to a concentration of 1 mg / mL, 2 μL of carboxypeptidase B (CpB) solution was added to 200 μL of the resulting solution at a concentration of 5 mg / mL. The mixture was then stirred and incubated at 37 °C for 2 hours. The test sample was dialyzed three times with water. During dialyzing, the test solution was placed in a 0.5 mL ultracentrifuge tube (Amicon) and centrifuged at 10,000 rpm for 10 minutes on a 5417R centrifuge (Eppendorf). After measuring the absorbance of the solution relative to water using a Cary 50 biospectrophotometer, a series of test probes at a concentration of 2 mg / mL were prepared. In a 96-well plate (Bio-Rad), 3 μL of labeling buffer (from the HT protein charge isomer labeling kit) was added to each well, 15 μL of test solution was added to each well, and 3 μL of staining mixture (from the HT protein charge isomer labeling kit) was added to each well. After the plate was placed in a dark environment for 10 minutes, water was added to each well at a rate of 36 μL / well, and the mixture was stirred by pipetting. The solution was then centrifuged at 1000 rpm in a 5417R centrifuge (Eppendorf).

[0635] Working solution preparation and chip filling

[0636] Working solutions and chips were prepared using the HT protein charge isomer labeling kit according to the manufacturer's instructions. Related analyses were performed according to standard procedures, using the "HT protein charge isomer 90s" assay.

[0637] Example 9: Determination of sample purity under reducing and non-reducing conditions using Labchip GXII (Caliper)

[0638] Test sample preparation

[0639] Denaturing and reducing solutions were prepared simultaneously using 700 μL of HT protein expression sample buffer. The reduced sample was prepared by adding 24.5 μL of 1M dithiothreitol (DTT), while the non-reduced sample was prepared by adding 24.5 μL of 1M iodoacetamide (IAM) as an alkylating agent to the sample buffer.

[0640] For each sample, prepare two microtubes as follows: one tube contains 35 μL of denaturing buffer; the other contains 35 μL of reducing buffer. After diluting the sample to a concentration of 2 mg / mL, add 5 μL of sample to each pair of tubes. Then, denature the sample at 100 °C for 5 minutes. After vortexing each tube, add 70 μL of water per tube, and then continue vortexing. Transfer 44 μL of each sample to the wells of a 96-well plate.

[0641] Working solution preparation and chip filling

[0642] Working solutions and chips were prepared using the HT protein expression kit according to the manufacturer's instructions. Related analyses were performed according to standard procedures, using the "HT Protein Expression 200" analytical method.

[0643] Example 10: Determination of the charge distribution characteristics of a sample by ion exchange (IE) high performance liquid chromatography (HPLC)

[0644] Chromatographic column: ProPac WCX-10 analytical column, 4×250mm;

[0645] Pre-column: Pro Pac WCX-10G, 4×50mm;

[0646] Column temperature: 30℃;

[0647] Mobile phase rate: 0.7 mL / min;

[0648] Injection volume: 50 μL;

[0649] Sample concentration: 1 mg / mL;

[0650] Detection wavelength: 220nm;

[0651] Wash-off time: 60 minutes;

[0652] Mobile phase:

[0653] Elution buffer A: 0.03M 2-(N-morpholino)ethanesulfonic acid (MES), pH = 6.0.

[0654] Elution buffer B: 0.03 M M EES, 0.5 M NaCl, pH = 6.0

[0655] Elution buffer A gradient: 86%–0%–86%.

[0656] Before analysis, the test samples were treated with carboxypeptidase at +37℃±1°C for 2 hours.

[0657] The absolute change in the charge distribution characteristics after treatment is calculated according to the following formula:

[0658] Δ = |Acid fraction content before treatment - Acid fraction content after treatment| + |Main fraction content before treatment - Main fraction content after treatment| + |Alkali fraction content before treatment - Alkali fraction content after treatment|.

[0659] Example 11: Determination of low molecular weight impurities by vertical electrophoresis (VEP) of reduced and non-reduced polyacrylamide gel (PAG)

[0660] Polyacrylamide gel (PAAG) was prepared between glass plates in the presence of sodium dodecyl sulfate, comprising a 4% PAAG concentration layer and a 12.5% ​​PAAG (reducing conditions) / 8% PAAG (non-reducing conditions) separation layer.

[0661] After assembling and installing the electrophoresis tank according to the instructions for the vertical electrophoresis apparatus, the probe was prepared by diluting the sample with purified water to a final concentration of 1 mg / mL. 40 μg volume equivalents were taken, and the prepared test sample probe was mixed with 4× sample buffer containing 2-mercaptoethanol (reduced) and without 2-mercaptoethanol (non-reduced) at a ratio of 3:1 (volume / volume), and then stirred. The resulting solutions were incubated at (99±1)°C for 3 minutes (for samples containing 2-mercaptoethanol) and at (99±1)°C for 1 minute (for samples without 2-mercaptoethanol), respectively. After the solutions cooled to room temperature, they were mixed and added to the PAG loading wells below the electrode buffer layer.

[0662] Subsequently, constant current electrophoresis was performed using a water-cooling system. The power supply parameters were set as follows: first, a voltage of 110V was applied to allow the dye front to pass through the stacking gel; when the dye front moved to the lower separating gel at a height of 5-7mm, the voltage was increased to 180V; when the dye front reached the bottom of the gel, the power was turned off.

[0663] After electrophoresis, the gel was peeled off the glass plate, and the proteins were fixed in fixative at room temperature for 16–18 hours. Subsequently, the gel was stained (Acid Blue 83 solution), washed, and clear bands were obtained. After gel scanning, the purity and impurity content of the test samples were evaluated using GelPro software.

[0664] Example 12: Relative activity determination

[0665] The relative activity of monoclonal anti-PD-1 antibodies was evaluated by assessing their ability to specifically bind to PD-1 protein on the surface of Jurkat-PD-1-NFAT cell membranes. One day prior to analysis, PDL-1 was immobilized on the walls of culture plates. The next day, after washing the plates, 50 μL of phytohemagglutinin P solution (PanEco, Russia, catalog number: M021) was added to each well. Subsequently, serial dilutions of standards and test samples were prepared using the Freedom Evo robot and added to each well at a rate of 10 μL. Jurkat-PD-1-NFAT cell suspension was added to each well at a rate of 40 μL, and the plates were incubated at 37°C with 5% CO2 for 4–6 hours. All steps were performed under aseptic conditions.

[0666] After incubation, BioGlo (Promega, USA, catalog number: G7941) solution was added at a rate of 100 μL / well, and the luminescence level was measured.

[0667] Using Magellan software, a four-parameter curve relating the average luminescence value to protein concentration was constructed for the standard and test sample solutions located on the same plate.

[0668] The relative specific activity (%, relative potency) of the test sample is calculated using the following formula:

[0669]

[0670] Among them, ED 50 st: the half-maximal effective dose (ng / mL) of the standard;

[0671] ED 50 test: The half-maximal effective dose (ng / mL) of the test sample.

[0672] Calculate the average relative activity from three independent measurements (three different culture plates) and use this value as the final result.

[0673] Example 13: Viscosity Measurement

[0674] The dynamic viscosity of the test solution was measured using a CAP2000+L (Brookfield) viscometer according to the rotational viscosity measurement method.

[0675] Example 14: Study on the Source of Buffer Solution

[0676] Test formula

[0677] This study selected four buffer solutions and chose the molar concentration and pH value of the buffer solution based on the limitations of possible subcutaneous administration and the antibody RI (the selection of the test solution RI value was based on the minimum possible physiological value).

[0678] Composition (per 1 mL):

[0679]

[0680] Determining Colloidal Stability Through PEG Aggregation

[0681] In this study, each sample was measured in triplicate. The results are shown in Table 1 and... Figure 1 .

[0682] Table 1: Average absorbance of the freshly prepared solution at 400 nm wavelength

[0683]

[0684] The results show that the samples exhibited the highest colloidal stability to PEG in histidine buffer and acetate buffer. The aggregation of the phosphate and citrate compositions at 6% PEG indicates that their colloidal stability was unsatisfactory, therefore these two compositions were excluded from subsequent studies. Based on this result, acetate buffer and histidine buffer were selected for subsequent pH / molar concentration selection.

[0685] Example 15: Selection of Solution pH / Buffering Capacity

[0686] Test formulation (per mL)

[0687]

[0688]

[0689] The study was conducted under thermal stress at 50°C for 72 hours, and the results are shown in Table 2. Sample homogeneity was determined by SEHPLC and electrophoresis performed using a Labchip system, while charge distribution characteristics were analyzed using a Labchip system.

[0690] Table 2: Summary of Quality Indicators Before and After Thermal Stress Treatment

[0691]

[0692]

[0693]

[0694] *The change is calculated using the following formula: Δ = Content of fraction after treatment - Content of fraction before treatment Best result -Worst Result -Initial reference or average result

[0695] The results of this study indicate that the presence of monoclonal anti-PD-1 antibody (Prolgolimab) in aqueous solution increases the pH level. A 20 mM buffer solution exhibits the greatest pH stabilizing ability.

[0696] According to SE HPLC results, the impurity increase of all tested samples after thermal stress treatment was no greater than 0.5%, thus exhibiting high aggregation stability. Histidine-based samples with pH = 5.5–6.0 and acetate-based samples with pH = 5.0–5.5 showed the best stability.

[0697] All formulations exhibited similar stability in terms of charge distribution characteristics, and the quantitative differences in fraction content variations among different formulations were all within the upper and lower limits of the accuracy of the respective methods.

[0698] According to the results of reducing gel electrophoresis, all histidine-based formulations showed higher stability, while under non-reducing conditions, the acetate-based solution yielded the best results.

[0699] Based on the data regarding the pH, purity, and charged form distribution characteristics of monoclonal anti-PD-1 antibodies, the following excipient formulations are recommended for further research:

[0700]

[0701] Example 16: Selection of Histidine-based Pharmaceutical Compositions

[0702] Test formula

[0703] When screening stable pharmaceutical compositions based on a 20 mM histidine buffer solution at pH 5.5, the following excipients were selected: mannitol; trehalose dihydrate; and sucrose (penetrating agent). All test solutions were isotonic.

[0704] Test sample (mg / 1mL)

[0705]

[0706]

[0707] Sample stability was evaluated using the following tests: 72 hours of thermal stress treatment at 50°C; 120 hours of shaking at 800 rpm; and a single freezing at -16 to -20°C followed by thawing at +25 ± 1°C. Solution turbidity was evaluated using 400 nm spectrophotometry. Sample homogeneity was determined by SE HPLC and electrophoresis using a Labchip system. Charge distribution characteristics were analyzed using a Labchip system.

[0708] Table 3: Summary of the quality index results of the samples before and after treatment obtained by gel filtration and ultraviolet spectrophotometry.

[0709]

[0710]

[0711] -Best Result -Worst Result - Initial reference or average results Table 4: Summary of the study results on the acid-base distribution characteristics of samples before and after treatment

[0712]

[0713]

[0714] *The change is calculated using the following formula: Δ = (After fractional content treatment - Before fractional content treatment)

[0715] **The absolute change is calculated using the following formula: Δ=|Acid fraction content before treatment - Acid fraction content after treatment| +|Alkali fraction content before treatment - Alkali fraction content after treatment| +|Main fraction content before treatment - Main fraction content after treatment|

[0716] -Best Result -Worst Result -Initial reference or average result

[0717] Table 3 shows that mannitol negatively impacts the thermal and colloidal stability of monoclonal anti-PD-1 antibodies in 20 mM histidine buffer solution at pH 5.5: during 96 hours of thermal stress treatment, the increase in impurities, as determined by SE HPLC, ranged from 1.24% to 3.18%; and during the 120-hour shaking test, visible aggregation occurred in the solution. Furthermore, the mannitol-containing formulation also exhibited a negative effect on stability during low-temperature concentration: after one round of freezing / thawing, the increase in impurities, as determined by SE HPLC, ranged from 1.05% to 4.45%, significantly higher than other formulations.

[0718] The formulation containing L-proline also exhibited visible aggregation during shaking tests, indicating low colloidal stability. After freezing / thawing and heat stress treatment, the increase in impurities in the L-proline-containing formulation was, on average, greater than 1%, as determined by SE HPLC.

[0719] The experiment revealed that Prolgolimab exhibits high thermal and colloidal stability in formulations containing trehalose dihydrate but without solubilizers such as polysorbate 80 and poloxamer 188 (no significant changes in quality indicators were observed under all treatments).

[0720] The following components exhibit optimal stabilizing effects on monoclonal anti-PD-1 antibodies in 20 mM histidine buffer: trehalose dihydrate, sucrose, and their respective combinations with glycine. These formulations can be used in hydrophilic formulations of monoclonal antibodies.

[0721] Stability evaluation of histidine-based candidate formulations after lyophilization

[0722] For the samples that showed the best stability in the previous screening stage, testing was conducted to verify whether they could be made into lyophilized products suitable for preparing infusion solutions.

[0723] In this process, a solution of the monoclonal anti-PD-1 antibody Prolgolimab at a concentration of 20 mg / mL or 100 mg / mL was placed in a Class I hydrolyzed glass vial, which was then loosely capped with a grooved rubber stopper. The vial containing the solution was placed in a lyophilization chamber for automated lyophilization. The freezing temperature was -40°C, and the pressure for the first drying step was (0.10 ± 0.03) mbar. For the second drying step, the temperature was increased, and the pressure was set to (0.05 ± 0.02) mbar. After the drying step, the pressure was reduced to -0.76 ± 0.03 mbar for vacuum treatment. The vial was then capped with a rubber stopper, the vacuum was released, and the pressure was restored to atmospheric pressure. The capped vials were used as lyophilized samples of the monoclonal anti-PD-1 antibody Prolgolimab in subsequent studies and stored at 2–8°C.

[0724] To verify protein stability, the samples were rehydrated after lyophilization. The monomer content was evaluated using SE HPLC, and the charge distribution characteristics were analyzed using a Labchip system. Furthermore, the pH values ​​before and after lyophilization were measured. The results are shown in Table 5.

[0725] During the lyophilization and rehydration of the selected histidine-based formulations, minor pH changes were observed, all within the upper and lower limits of the accuracy of the respective methods. SE HPLC and charge distribution analysis after rehydration showed that all formulations exhibited high stability in terms of purity. However, LabChip (Caliper) gel electrophoresis analysis revealed significant differences in purity among different samples. The samples with the highest stability in this parameter were formulations 22, 27, and 36, which contained trehalose dihydrate. The formulations containing L-proline showed significant changes in charge distribution characteristics and were therefore not recommended. Therefore, the results demonstrate that formulations 22 and 27, with a protein concentration of 100 mg / mL, are suitable for lyophilization.

[0726] Table 5: Summary of quality index results before and after lyophilization of the formulation based on 20mM histidine buffer solution at pH=5.5

[0727]

[0728]

[0729] *The change is calculated using the following formula: Δ = (After fractional content treatment - Before fractional content treatment)

[0730] **The absolute change is calculated using the following formula: Δ=|Acid fraction content before treatment - Acid fraction content after treatment| +|Alkali fraction content before treatment - Alkali fraction content after treatment| +|Main fraction content before treatment - Main fraction content after treatment|

[0731] -Best Result -Worst Result -Initial reference / average result

[0732] Preparation of high-concentration formulations and confirmation of stability under accelerated aging conditions.

[0733] Based on the screening results for liquid and hydrophilic formulations, the following formulations were selected for stability studies under accelerated aging conditions:

[0734]

[0735] For solutions containing high concentrations of monoclonal anti-PD-1 antibody, the trehalose dihydrate content was reduced to equilibrate the osmotic pressure to physiological levels (approximately 300 mOsm / kg) and decrease the viscosity to below 100 cP. In this study, the stability of the samples was investigated by accelerated aging at +37°C, and the samples were analyzed by SE HPLC, IE HPLC, and VEP. Furthermore, the relative specific activity of Prolgolimab was determined. The results are shown in Tables 6 and 7.

[0736] Table 6: Stability analysis results at 37℃

[0737]

[0738]

[0739] Table 7: Stability analysis results at 37℃

[0740]

[0741]

[0742] Example 17: Selection of Acetate-Based Pharmaceutical Compositions

[0743] Test formula

[0744] When screening stable pharmaceutical compositions based on a 20 mM acetate buffer solution at pH 5.0, the following excipients were selected: mannitol; trehalose dihydrate; sucrose (penetrating agent); L-proline (penetrating agent and stabilizer); glycine (penetrating agent and stabilizer); polysorbate 80 and poloxamer P188 (solvents). All test solutions were isotonic.

[0745] Test formulation (per mL)

[0746]

[0747] Sample stability was evaluated using the following tests: 72 hours of thermal stress treatment at 50°C; 120 hours of shaking at 800 rpm; and a single freezing at -16 to -20°C followed by thawing at +25±1°C. Solution turbidity was evaluated using 400 nm spectrophotometry. Sample homogeneity was determined by SE HPLC and electrophoresis using a Labchip system. Charge distribution characteristics were analyzed using a Labchip system.

[0748] Table 8: Results of Sample Gel Filtration Method / Ultraviolet Spectrophotometry

[0749]

[0750] -Best Result -Worst Result - Initial control or average results

[0751] Table 9: Results of acid-base distribution characteristics analysis of samples

[0752]

[0753] *The change is calculated using the following formula: Δ = (After fractional content treatment - Before fractional content treatment)

[0754] **The absolute change is calculated using the following formula: Δ=|Acid fraction content before treatment - Acid fraction content after treatment| +|Alkali fraction content before treatment - Alkali fraction content after treatment| +|Main fraction content before treatment - Main fraction content after treatment|

[0755] -Best Result -Worst Result - Initial control or average results

[0756] The results of this study indicate that mannitol negatively impacts the thermal and colloidal stability of monoclonal anti-PD-1 antibodies in 20 mM acetate buffer solution at pH 5.0: during 96 hours of thermal stress treatment, the increase in impurities, as determined by SE HPLC, ranged from 0.48% to 2.59%; during the 120-hour shaking test, no visible aggregation occurred in any of the test solutions (see UV spectrophotometric results). Furthermore, the mannitol-containing formulation also exhibited a negative effect on stability during low-temperature concentration: after one round of freezing / thawing, the increase in impurities, as determined by SE HPLC, ranged from 0.69% to 9.44%, significantly higher than other formulations.

[0757] Formulations containing L-proline also exhibited lower thermal stability in terms of purity as determined by SE HPLC analysis: the increase in impurities after 96 hours of treatment was 1.64–1.93%. After freezing / thawing, the increase in impurities in L-proline-containing formulations was not higher than the average of most high-stability formulations in the group. During the experiments, solubilizers did not show any effect on the thermal or colloidal stability of the protein. In formulations containing trehalose dihydrate, Prolgolimab exhibited high stability in all treatment experiments when surfactants such as polysorbate 80 or poloxamer 188 were absent. The following substances showed the best stabilizing effect on monoclonal anti-PD-1 antibodies in 20 mM acetate buffer: trehalose dihydrate, sucrose, and their respective combinations with glycine. These formulations can be used for hydrophilic formulations of monoclonal antibodies.

[0758] Stability evaluation of candidate formulations based on acetate after lyophilization

[0759] For the samples that showed the best stability in the previous screening stage, testing was conducted to verify whether they could be made into lyophilized products suitable for preparing infusion solutions.

[0760] In this process, a solution of the monoclonal anti-PD-1 antibody Prolgolimab at a concentration of 20 mg / mL or 100 mg / mL was placed in a Class I hydrolyzed glass vial, which was then loosely capped with a grooved rubber stopper. The vial containing the solution was placed in a lyophilization chamber for automated lyophilization. The freezing temperature was -40°C, and the pressure for the first drying step was (0.10 ± 0.03) mbar. For the second drying step, the temperature was increased, and the pressure was set to (0.05 ± 0.02) mbar. After the drying step, the pressure was reduced to -0.76 ± 0.03 mbar for vacuum treatment. The vial was then capped with a rubber stopper, the vacuum was released, and the pressure was restored to atmospheric pressure. The capped vials were used as lyophilized samples of the monoclonal anti-PD-1 antibody Prolgolimab in subsequent studies and stored at 2–8°C.

[0761] To verify protein stability, the samples were rehydrated after lyophilization. The monomer content was evaluated using SE HPLC, and the charge distribution characteristics were analyzed using a Labchip system. Furthermore, the pH values ​​before and after lyophilization were measured. The results are shown in Table 8.

[0762] Significant pH changes ranging from 0.26 to 0.45 were detected during the lyophilization and rehydration of the lyophilized product using the selected acetate-based formulation. This change is likely due to acetate loss during lyophilization. Therefore, when the above situation is observed, the acetate-based formulation is the recommended formulation suitable for lyophilization.

[0763] After rehydration, all formulations exhibited high stability in terms of purity as measured by SE HPLC and charge distribution characterization. These results demonstrate that formulations 10 and 12, with a protein concentration of 100 mg / mL, are suitable for lyophilization.

[0764] Table 10: Stability results of the formulation based on 20 mM acetate buffer solution at pH 5.0 after lyophilization.

[0765]

[0766]

[0767] *The change is calculated using the following formula: Δ = (After fractional content treatment - Before fractional content treatment)

[0768] **The absolute change is calculated using the following formula: Δ=|Acid fraction content before treatment - Acid fraction content after treatment| +|Alkali fraction content before treatment - Alkali fraction content after treatment| +|Main fraction content before treatment - Main fraction content after treatment|

[0769] -Best Result -Worst Result - Preparation of high-concentration forms for initial control or average results and confirmation of stability under accelerated aging conditions

[0770] Based on the screening results for liquid and hydrophilic formulations, the following formulations were selected for stability studies under accelerated aging conditions:

[0771]

[0772] For solutions containing high concentrations of monoclonal anti-PD-1 antibody, the trehalose dihydrate content was reduced to equilibrate the osmotic pressure to physiological levels (approximately 300 mOsm / kg) and decrease the viscosity to below 100 cP. In this study, the samples were aged at +37°C and analyzed by SE HPLC, IE HPLC, and VEP. Furthermore, the relative specific activity of Prolgolimab was determined. The results are shown in Tables 9 and 10.

[0773] Table 11: Stability analysis results at 37℃

[0774]

[0775]

[0776] Table 12: Stability analysis results at 37℃

[0777]

[0778]

[0779] Although the present invention has been described above with reference to preferred embodiments, it will be understood that modifications that will be obvious to those skilled in the art can be made. Such modifications and alterations are considered to be within the spirit and scope of the present invention.

[0780] The foregoing has provided a detailed description of representative, non-limiting embodiments of the present invention. This detailed description is intended only to provide those skilled in the art with further details for practicing preferred aspects of the invention and is not intended to limit the scope of the invention. Furthermore, all other features and techniques disclosed above and below can be used alone or in combination with other features and techniques.

[0781] Furthermore, the combination of features and steps disclosed in the above detailed description and experimental embodiments is, in the broadest sense, not necessarily essential to the practice of the invention; rather, it is merely intended to specifically describe particular examples of the invention. Additionally, to further provide other useful embodiments of the invention, the various features of the above-described embodiments and the appended independent and dependent claims can be combined in ways not specifically and explicitly enumerated herein.

[0782] The following are other exemplary studies using various aqueous pharmaceutical compositions containing the anti-PD-1 antibody Prolgolimab for the treatment of malignancies such as: melanoma, including inoperable or metastatic melanoma, and early-stage melanoma before and after definitive treatment; and lung cancer, including non-small cell lung cancer (NSCLC), including inoperable or metastatic NSCLC. The exemplary pharmaceutical compositions used in these studies are characterized by the compositions shown below (Table 12.1).

[0783] Table 12.1I. Composition per 1 mL:

[0784]

[0785] II. Composition per 1 mL:

[0786]

[0787] Key information for clinical development of test products

[0788] In a Phase I dose-escalation clinical trial, the pharmacokinetics, pharmacodynamics, safety, and immunogenicity of BCD-100 (Prolgolimab) as monotherapy were investigated after intravenous administration. This study demonstrated the safety and benefit of the test product in patients with various common forms of malignancy (melanoma, NSCLC) at various sites. Based on the results of this study, two BCD-100 administration regimens were selected for further clinical development: 3 mg / kg intravenously every three weeks; and 1 mg / kg intravenously every two weeks.

[0789] The BCD-100-2 / MIRACULUM trial evaluated the pharmacokinetics, efficacy, safety, and immunogenicity of two monotherapy dosage regimens: 3 mg / kg intravenously every three weeks and 1 mg / kg intravenously every two weeks. An interim analysis was conducted after 6 months to assess the primary endpoint (ORR). Results showed that BCD-100 had a good safety profile at all dosage regimens, and both regimens demonstrated efficacy in patients with unresectable or metastatic melanoma: a disease control rate of approximately 60% and an ORR of approximately 30%.

[0790] Example 18: Use of Prolgolimab (BCD-100) in the treatment of patients with various common forms of malignancy in various sites (Phase I trial, BCD-100-1)

[0791] Experimental Design

[0792] The BCD-100-1 trial investigated the pharmacokinetics and tolerability of BCD-100 (Prolgolimab). The BCD-100-1 trial (NCT03050047) was a phase I, multicenter, unblinded trial in patients with solid tumors, conducted in the Russian Federation. The primary objective of this trial was to evaluate the pharmacokinetics and clinical pharmacology parameters of BCD-100. Key characteristics of the BCD-100-1 trial are shown in Table 13.

[0793] Table 13: Characteristics of the BCD-100-1 test

[0794]

[0795]

[0796] The first patient was initially administered an initial dose of BCD-100 (0.3 mg / kg) every two weeks. If no dose-limiting toxicity (DLT) was observed within 4 weeks, the patient continued to receive the medication every two weeks, but the dose was gradually increased to 1 mg / kg. Following the first patient, the trial continued using a traditional 3+3 dose-escalation design, meaning that if no DLT was observed, three patients were sequentially enrolled every four weeks at the next dose level. At each dose level, BCD-100 was administered for 85 days (approximately 3 months), or until signs of DLT / disease progression were observed.

[0797] If a DLT event is observed in one patient at a certain dose level, three patients in the next cohort receive the same dose level of the product. Thus, a total of six patients receive that dose of BCD-100. If a DLT event is observed in two or more patients at a certain dose level, dose escalation for a new cohort of patients is discontinued.

[0798] Patients treated with BCD-100 were followed up for 126 days. During this period, adverse events were monitored, medical examinations were performed, and blood analyses were conducted (once every two weeks for the first 28 days, and once every 28 days thereafter).

[0799] Test results

[0800] Summary of patient characteristics

[0801] This study included 15 patients aged 18 years and older with common forms of malignant solid tumors (including melanoma, choroidal melanoma, NSCLC, and renal cell carcinoma) at various sites, both male and female. Inclusion criteria were: ECOG score of 0–2; and at least one measurable target lesion (excluding bone metastases) meeting RECIST 1.1 criteria. Patient disease characteristics and distribution are shown in Tables 14 and 15.

[0802] Table 14: Patient Disease Characteristics (BCD-100-1 Trial)

[0803]

[0804] Table 15: Patient Grouping (BCD-100-1 Trial)

[0805]

[0806] The final analysis included data from 15 patients (6 patients in the BCD-100 1 mg / kg cohort (1 of whom received dose escalation as described above), 6 patients in the BCD-100 3 mg / kg cohort, and 3 patients in the BCD-100 10 mg / kg cohort). All patients received treatment during the main trial phase (85 days).

[0807] All patient cohorts were balanced in terms of key demographic characteristics. Patients were evenly distributed by sex, with 53.33% being female (8 patients) and 46.67% being male (7 patients). The median age of the patients was 56 years (minimum age 35 years, maximum age 77 years).

[0808] Regarding the primary disease characteristics, melanoma was the most common (9 out of 15 patients). In addition to melanoma patients, there were 4 NSCLC patients, 1 pleural mesothelioma patient, and 1 renal cell carcinoma patient. At inclusion, the median duration of illness for all patients was 17.07 months, with a minimum of 0.6 months and a maximum of 91.2 months.

[0809] Summary of clinical safety results from Phase I trials

[0810] A study of the safety of the study in all patients (n=15) receiving at least one dose of the study product revealed adverse events and / or serious adverse events in all 15 patients, totaling 247 events. Statistical analysis showed no significant differences in adverse events between the cohorts (p=0.567, Kruskal-Wallis test). A summary of product safety data is shown in Table 16 below.

[0811] 40.00% of patients (6 out of 15) experienced adverse events of severity grade 3 as defined in CTCAE 4.03, meaning two patients in each cohort. Researchers considered the following four patients with grade 3 or higher adverse events to be treatment-related: one patient receiving BCD-100 at a dose of 1 mg / kg; one patient receiving BCD-100 at a dose of 3 mg / kg; and two patients receiving BCD-100 at a dose of 10 mg / kg. The majority of these adverse events were hematological.

[0812] Serious adverse events occurred in three patients who received a dose of 1 mg / kg BCD-100 and in one patient who received a dose of 3 mg / kg BCD-100.

[0813] During the trial, one dose-limiting toxicity (DLT) event occurred. Patient 13-09 developed endocrine abnormalities (decreased TSH) and autoimmune thyroid disease (an immune-mediated grade 2 adverse event as defined in CTCAE 4.03) after receiving two doses of 3 mg / kg BCD-100. The Data Monitoring and Safety Committee classified this event as a DLT. Patient 13-09 subsequently continued treatment in this study, and their condition was classified as "stable" according to RECIST 1.1 and irRC guidelines.

[0814] In addition to autoimmune thyroid disease, immune-mediated adverse events also occurred in 33.33% of patients (5 out of 15): 33.33% (2 out of 6) received BCD-100 at a dose of 1 mg / kg; 50.00% (3 out of 6) received BCD-100 at a dose of 3 mg / kg; and none of the patients receiving BCD-100 at a dose of 10 mg / kg experienced such adverse events. All of these adverse events were grade 1 severity events as defined in CTCAE 4.03 and were not identified as DLT events.

[0815] During the study, three patients temporarily discontinued treatment due to adverse events / serious adverse events: 16.67% (1 out of 6) of patients receiving BCD-100 at a dose of 1 mg / kg; and 33.33% (2 out of 6) of patients receiving BCD-100 at a dose of 3 mg / kg. All adverse events leading to temporary treatment discontinuation were immune-related events (Patient 10-03 receiving BCD-100 experienced grade 1 hyperthyroidism; Patient 13-10 experienced grade 1 autoimmune thyroid disease; and Patient 13-09 experienced grade 2 autoimmune thyroid disease (the latter two patients were receiving BCD-100 at a dose of 3 mg / kg)). Patient 10-03 discontinued treatment in this study, while patients 13-09 and 13-10 continued treatment in this study after receiving glucocorticoid therapy. All treatment interruptions were within the range of 2–4 weeks specified in the trial protocol.

[0816] In this study, three patients discontinued product administration due to disease progression (two after five doses of 1 mg / kg and one after five doses of 10 mg / kg). All these discontinuations were due to disease progression, not the administration of BCD-100.

[0817] In this study, one patient discontinued treatment due to a serious adverse event and subsequently died. Patient 13-10 experienced a grade 5 right hemisphere stroke after receiving three doses of the product at 3 mg / kg. Researchers believe this adverse event may be related to the treatment administered in this study.

[0818] No differences were observed between patient groups regarding changes in laboratory and physiological parameters over time. Although the analysis of these parameters revealed a few isolated cases of statistically significant differences, we consider these differences to be natural and normal variations. Deviations in biochemical blood tests and coagulation tests (elevated transaminase and bilirubin levels, deviations in coagulation indices, etc.) are mostly typical of this type of subject population (cancer patients unsuitable for surgery) and are predictable transient biases. No abnormalities were found after parameter normalization, requiring no further processing.

[0819] The above results demonstrate that the BCD-100 product has good safety at any of the intravenous administration doses studied.

[0820] Table 16: Safety Data (BCD-100-1 Test)

[0821]

[0822] Immunogenicity

[0823] Immunogenicity was assessed at patient enrollment, on day 28 of study treatment, and at the end of the main study phase (day 85). Additionally, for patients continuing treatment with the BCD-100 product, immunogenicity was subsequently assessed every 42 days. The immunogenicity assessment consisted of two steps: first, screening patient serum samples for the presence of binding antibodies (BABs); second, screening samples for the presence of neutralizing antibodies (NABs) using positive BAB titers.

[0824] Immunogenicity evaluation of serum samples from all available patients (n=15) at patient enrollment and in each subsequent analysis revealed that none of the samples contained BAB. Therefore, no NAT analysis was performed because none of the samples contained BAB.

[0825] Summary of Phase I Pharmacokinetic / Metabolism Results

[0826] Pharmacokinetic (PK) analysis included data from patients (n=15) whose serum samples were used to determine BCD-100 concentrations after the first administration of the product and who did not have more than three missed, lost, or discarded samples.

[0827] In the pharmacokinetic analysis, patients were divided into the following groups (after the first administration and after all subsequent administrations):

[0828] • First patient (BCD-100 dose of 0.3 mg / kg) (n = 1);

[0829] • First cohort of patients (BCD-100 dose of 1 mg / kg) (n = 5);

[0830] • Second cohort of patients (BCD-100 dose of 3 mg / kg) (n = 6);

[0831] • Third cohort of patients (BCD-100 dose of 10 mg / kg) (n=3).

[0832] • Pharmacokinetic analysis included an evaluation of the standard pharmacokinetic characteristics (distribution and excretion) of the study product after multiple intravenous administrations. The results are shown in Table 17.

[0833] • For pharmacokinetic analysis (for serum BCD-100 levels), blood samples were collected from all enrolled patients at each dose level. Sampling time points were as follows: before the first administration; 30 minutes, 2 hours (±15 minutes), 4 hours (±15 minutes), 6 hours (±15 minutes), 24 hours (±1 hour), 48 hours (±2 hours), 192 hours (±8 hours), and 336 hours (±8 hours) after the first administration (before the second administration); and before subsequent bi-weekly administrations. In addition to blood samples used to determine serum BCD-100 concentrations, samples were also collected for immunogenicity analysis (to evaluate the potential effect of the product antibody on the pharmacokinetic properties of BCD-100). Serum BCD-100 concentrations were determined using a validated ELISA method.

[0834] Table 17: Pharmacokinetic Parameters (BCD-100-1 Test)

[0835]

[0836] Except C min Apart from the calculated values ​​after 6 applications (12 weeks) of the product, all other parameters are measured values ​​after a single application of the product.

[0837] The following parameters showed statistically significant differences among the three cohorts receiving different doses: AUC (0-336h) (p = 0.0088, Kruskal-Wallis test); AUMC (0-336h) (p = 0.0088, Kruskal-Wallis test); AUC (0-∞) (p = 0.0123, Kruskal-Wallis test); C max(p = 0.0103, Kruskal-Wallis test). Pharmacokinetic characteristics were better at 3 mg / kg and 10 mg / kg doses compared to 1 mg / kg. However, pharmacokinetic, efficacy, and safety parameters did not show a correlation with dose.

[0838] Figure 1 and Figure 2 The figure shows the BCD-100 product concentration curves for all dose cohorts (1 mg / kg cohort [n=5], 3 mg / kg cohort [n=6], and 10 mg / kg cohort [n=3]) during the period from the first administration to week 10.

[0839] Calculations of pharmacokinetic parameters showed that BCD-100 concentration was directly proportional to the administered dose, peaking sometime between 30 minutes and 6 hours after administration, and then gradually decreasing. Based on these results, there was no correlation between the half-life and the amount of product administered. 1 / 2 The parameter values ​​are typical for monoclonal antibodies (12–18 days).

[0840] Summary of Phase I Pharmacodynamic Results

[0841] The pharmacodynamic evaluation subjects were all patient serum samples (n=15) available at the time of patient selection and in each subsequent analysis.

[0842] In the pharmacodynamic study, patients were divided into the following groups:

[0843] • First patient (BCD-100 dose of 0.3 mg / kg) (n = 1);

[0844] • First cohort of patients (BCD-100 dose of 1 mg / kg) (n = 5);

[0845] • Second cohort of patients (BCD-100 dose of 3 mg / kg) (n = 6);

[0846] • Third cohort of patients (BCD-100 dose of 10 mg / kg) (n=3).

[0847] The saturation level of the PD-1 receptor under the influence of the BCD-100 product helps evaluate the interaction between the study product and its target, and given that PD-1 receptor blockade can activate anti-tumor immune responses, it is the ultimate indicator of the activity of monoclonal anti-PD-1 antibodies. For the pharmacodynamic evaluation (percentage saturation of the PD-1 receptor under the influence of the BCD-100 product), blood samples were collected from all patients. Sampling times for the pharmacodynamic evaluation were: before product administration; 4 hours and 336 hours after administration (but before the second administration); and before the sixth administration.

[0848] The results showed that PD-1 receptors were highly saturated (95%–100%) with BCD-100 product at all dose levels and in all cell populations (Table 18).

[0849] Table 18: Saturation of PD-1 receptors (BCD-100-1 assay).

[0850]

[0851]

[0852] Summary of Phase I clinical efficacy results

[0853] Of the 15 patients who received BCD-100, 14 were included in the efficacy evaluation. One patient discontinued treatment due to a serious adverse event (death). The tumor response to treatment was not evaluated by CT prior to the patient's death.

[0854] On day 85 after initiation of BCD-100 treatment, the overall response rate (partial response rate + complete response rate) and disease control rate (disease stabilization rate + partial response rate + complete response rate) were determined. Efficacy evaluation on day 85 after initiation of BCD-100 treatment was based on analysis of CT scan results. Tumor response was evaluated according to RECIST 1.1 and immune-related RECIST (irRECIST).

[0855] According to the irRECIST criteria for immunotherapy, disease control was 28.57% (4 out of 14 patients). The overall response rate was 7.14% (1 out of 14 patients). There were no significant differences in response rates between different patient cohorts (Table 19).

[0856] Table 19: Tumor Response (BCD-100-1 Test)

[0857]

[0858]

[0859] in conclusion

[0860] The analysis of the obtained data clearly demonstrates that BCD-100 has a good safety profile at various doses ranging from 0.03 mg / kg to 10.0 mg / kg. The following two administration regimens of BCD-100 appear to be the optimal choices for further clinical studies: 1 mg / kg every two weeks; and 3 mg / kg every three weeks.

[0861] Example 19: Use of Prolgolimab (BCD-100) in the treatment of patients with unresectable / metastatic melanoma (Phase II trial, BCD-100-2 / MIRACULUM)

[0862] Experimental Design

[0863] The trial, designated BCD-100-2 / MIRACULUM (MIRACULUM, NCT03269565), is a multicenter, non-blinded, randomized trial aimed at evaluating the pharmacokinetics, efficacy, safety, and immunogenicity of BCD-100 (Prolgolimab) monotherapy in previously untreated or previously treated patients with unresectable / metastatic melanoma. Specifically, the study compared a 3 mg / kg intravenous dose of BCD-100 administered every three weeks with a 1 mg / kg intravenous dose administered every two weeks. The study is being conducted in the Russian Federation and Belarus and is still ongoing. Data after one year of treatment have been obtained and analyzed.

[0864] Each efficacy parameter was evaluated individually for each group. The primary endpoint of efficacy evaluation was the overall response rate (partial response rate + complete response rate) of the subjects in this study relative to the background level of BCD-100 product treatment, as measured by irRECIST. The optimal treatment response at these stages was considered in the calculation of ORR (overall response rate) and disease control. Secondary endpoints of efficacy evaluation included progression-free survival and overall survival at 12 months after treatment initiation, disease control rate (disease stabilization rate + partial response rate + complete response rate), time to treatment response, and duration of treatment response. Figure 3 (Table 20).

[0865] Table 20: Characteristics of the BCD-100-2 / MIRACULUM (NCT03269565) test

[0866]

[0867] Test results

[0868] Summary of patient characteristics

[0869] A total of 131 patients, aged 18 years and older, were included. The patients were both male and female, and all had common melanomas, including choroidal melanoma. Some patients withdrew before the first dose of BCD-100 or discontinued the trial due to protocol violations; these were subsequently added. Ultimately, 126 patients received at least one dose of BCD-100 in the modified ITT program. Patient inclusion criteria were: an ECOG score of 0–1; and at least one measurable target lesion (excluding bone metastases) meeting RECIST 1.1 criteria. Figure 4 (Table 21).

[0870] Table 21: Patient Disease Characteristics (BCD-100-2 / MIRACULUM Trial)

[0871]

[0872]

[0873]

[0874] The safety analysis included all patients who received at least one administration of the study product (mITT (modified Intent-to-Treatment) patients (n=126)).

[0875] In the efficacy analysis, patients were divided into the following two groups:

[0876] • All patients who received at least one administration of the study product (mITT patients, n=126);

[0877] • All patients who received at least one administration of the study product and at least one planned dynamic CT scan for evaluating response (“protocol-compliant” patients, n=114).

[0878] • Pharmacokinetic analysis includes data from patients (n=125) who received at least one dose of BCD-100 product and whose regimen version was valid at the time of assay and for whom no more than three samples were missed, lost, or discarded.

[0879] Summary of clinical efficacy results from the BCD-100-2 / MIRACULUM trial

[0880] The evaluation results of the primary efficacy endpoint (ORR) showed that both dosage regimens of BCD-100 were adequately effective in all subjects. Specifically, the ORR and disease control rate were 40.68% and 67.80% in the first treatment group (PP patients), and 32.73% and 52.73% in the second group. The efficacy results in mITT patients were similar. Therefore, both treatment groups achieved the expected target response rate (28%) and the cutoff value r (11 responses).

[0881] Analysis of different groups who had previously received various treatments showed that the minimum dose of BCD-100 at 1 mg / kg was highly effective in previously untreated patients (ORR of 50.00%).

[0882] Analysis of secondary efficacy endpoints confirmed that both doses of BCD-100 were sufficiently effective. Progression-free survival (PFS), overall survival (OS), and duration of response were comparable to those of top-performing PD-1 inhibitors.

[0883] The 12-month progression-free survival rate was 41.27% in the BCD-100 1 mg / kg group and 34.92% in the BCD-100 3 mg / kg group. The median progression-free survival time was 5.78 months (95% CI: 3.52–-) in the BCD-100 1 mg / kg group and 2.33 months (95% CI: 2.07–10.25) in the BCD-100 3 mg / kg group (p = 0.400, log-rank test). A detailed analysis of progression-free survival in the groups with different prior treatment histories (mITT patients according to irRECIST guidelines) revealed no statistically significant differences. The BCD-100 product is equally effective in both previously treated and previously treated patients.

[0884] Median overall survival (95% CI: -) in the BCD-100 1 mg / kg group 1The median overall survival (OS) was not reached at 13.8 months (95% CI: 13.2–14.7). The 12-month OS was 74.60% in the BCD-100 1 mg / kg group. The median OS was 15 months (95% CI: 9.99–-) in the BCD-100 3 mg / kg group, with a median observed survival of 14.5 months (95% CI: 13.9–15.2). The 12-month OS was 53.97% in the BCD-100 3 mg / kg group. No statistically significant differences in OS were found among the groups receiving different prior treatments. BCD-100 is equally effective in both previously treated and previously treated patients. No group in the 1 mg / kg BCD-100 group achieved the median OS (p = 0.800, log-rank test). In the BCD-100 3 mg / kg group (p = 0.900; log-rank test), the median overall survival was 16.8 months (95% CI: 9.33 to -) for previously untreated patients and 15 months (95% CI: 7.46 to -) for previously treated patients (Tables 22, 23, 24, 25). Figure 5 , Figure 6 , Figure 7 , Figure 8 ).

[0885] Table 221: Primary endpoints reflecting the efficacy of the product in PP patients (patients receiving treatment according to the protocol) (BCD-100-2 / MIRACULUM trial)

[0886]

[0887] Summary of clinical safety results of the BCD-100-2 / MIRACULUM trial

[0888] Both dosage regimens of BCD-100 demonstrated good safety (Table 23).

[0889] Table 23: Safety of BCD-100 Products (BCD-100-2 / MIRACULUM Test)

[0890]

[0891]

[0892] Based on the data above, the results obtained in this study are not contradictory to the known safety data of monoclonal anti-PD-1 receptor antibody products. Apart from the number of serious adverse events leading to death (significantly higher in the BCD-100 3 mg / kg group than in the other group), no statistically significant differences in product safety were found between the different dosages in this study.

[0893] Immunogenicity

[0894] Immunogenicity was evaluated from serum samples of all patients (n=121) available at patient enrollment and in each subsequent analysis. No patients had detectable BCD-100 BAB.

[0895] Summary of pharmacokinetic and metabolic results of the BCD-100-2 / MIRACULUM trial

[0896] This analysis showed that after a single intravenous administration of BCD-100 at a dose of 1–3 mg / kg, the concentration of the study substance in plasma initially increased linearly, followed by a uniform decrease. The half-life of the product was independent of the dose administered to the organism, and the characteristic time was 11.5–17 days, which is the typical half-life of IgG immunoglobulin. Figure 9 , Figure 10 ).

[0897] In subsequent administrations, the plasma concentration of BCD-100 continued to increase with increasing dose (according to C...). max and AUC 0-t,SS The parameters are consistent with the pharmacokinetic data of other anti-PD-1 antibody products reported in the literature.

[0898] Results from the clinical trial BCD-100-2 / MIRACULUM demonstrated that, relative to the background value during treatment, C min The levels remained consistently high, demonstrating that both the bi-weekly 1 mg / kg and bi-weekly 3 mg / kg dosage regimens of BCD-100 maintained therapeutic concentrations. Therefore, from a pharmacokinetic perspective, both BCD-100 dosage regimens were appropriate.

[0899] Because some patients lack a typical terminal elimination phase after a single administration, it is difficult to conduct comprehensive evaluations, including comparisons between different treatment groups. 1 / 2 Parameter evaluation.

[0900] Overall, the pharmacokinetic characteristics of BCD-100 product, administered at a dose of 1 mg / kg every two weeks and 3 mg / kg every three weeks, are characterized by a long residence time in the body sufficient to maintain stable therapeutic concentrations relative to a background value from multiple administrations. No dose-related correlations were observed in the study.

[0901] Summary of Pharmacodynamic Results from Phase I Trials

[0902] There were no differences in the percentage of PD-1 receptor saturation across all leukocyte subsets under the influence of BCD-100 product between the different treatment groups. Among patients analyzed for PD-1 saturation, the proportion of activated helper T cells and cytotoxic leukocytes with PD-1 saturation >99% was 33.33% (14 out of 42 patients). No statistically significant differences were found between the following treatment groups: 8 patients receiving BCD-100 1 mg / kg; and 6 patients receiving BCD-100 3 mg / kg.

[0903] Ki-67 is present only in cells undergoing mitosis and degrades within 1.5–2 hours after division, thus serving as a universal marker of cell proliferation. Ki-67 can be detected at telomeres, centromeres, and the nucleus. During treatment, an increase in the percentage of Ki-67-positive cytotoxic T cells was detected in both groups, but no significant difference was found. Furthermore, analysis of the Ki-67-positive cytotoxic T cell subset also revealed no statistically significant differences between the groups.

[0904] During treatment, the increase in the percentage of Th9 cells in the total helper T cell population was mainly observed in group 2 (BCD-100, 3 mg / kg, once every three weeks), but no significant difference was found. Furthermore, no significant differences were found when comparing across groups.

[0905] Based on existing literature, an initial increase in Th9 levels is associated with improved response to nivolumab in melanoma treatment. 3 It should be noted that this result was observed in only a limited number of patients (n=42), and these patients varied in terms of gender and major disease characteristics.

[0906] A comparative analysis of the Th9 component of the total helper T cell population in two groups exhibiting different types of responses to BCD-100 product treatment revealed no statistically significant differences between the two groups. However, the corresponding figure ( Figure 11 , Figure 12 This reflects a trend that patients with initially increased Th9 levels respond better to treatment.

[0907] in conclusion

[0908] The data obtained on the efficacy, safety, and pharmacokinetic characteristics of BCD-100 are sufficient to demonstrate the rationality of the two dosage regimens: 1 mg / kg once every two weeks and 3 mg / kg once every three weeks.

[0909] The international, multicenter, unblinded, randomized phase II trial BCD-100-2 / MIRACULUM in patients with unresectable / metastatic melanoma demonstrated a significant therapeutic advantage over known chemotherapy data, with results comparable to current best-in-class treatments. Considering the efficacy data and favorable safety profile obtained from the aforementioned trial, the use of BCD-100 in routine clinical practice can meet the benefit / risk balance for specific patient populations.

Claims

1. An aqueous pharmaceutical composition for an anti-PD-1 antibody, comprising the following: (a) Prolgolimab at a concentration of 15 mg / mL to 40 mg / mL as an antibody; (b) Trehalose dihydrate at a concentration of 95 mg / mL to 105 mg / mL; (c) Sodium acetate trihydrate at concentrations of 1.6 mg / mL to 1.9 mg / mL; and (d) Add acetic acid to a pH of 4.5-5.

5.

2. The aqueous pharmaceutical composition according to claim 1, characterized in that: The concentration of Prolgolimab was 20 mg / mL.

3. The aqueous pharmaceutical composition according to claim 1, characterized in that: The concentration of trehalose dihydrate is 100 mg / mL.

4. The aqueous pharmaceutical composition according to claim 1, characterized in that: The concentration of sodium acetate trihydrate is 1.7 mg / mL to 1.8 mg / mL.

5. The aqueous pharmaceutical composition according to claim 4, characterized in that: The concentration of sodium acetate trihydrate is 1.742 mg / mL.

6. The aqueous pharmaceutical composition according to any one of claims 1 to 2, characterized in that: Add acetic acid to bring the pH to 5.

0.

7. An aqueous pharmaceutical composition for an anti-PD-1 antibody, comprising the following: (a) Prolgolimab at a concentration of 90 mg / mL to 110 mg / mL as an antibody; (b) Trehalose dihydrate at a concentration of 75 mg / mL to 85 mg / mL; (c) Sodium acetate trihydrate at concentrations of 1.6 mg / mL to 1.9 mg / mL; and (d) Add acetic acid to a pH of 4.5-5.

5.

8. The aqueous pharmaceutical composition according to claim 7, characterized in that: The concentration of Prolgolimab was 100 mg / mL.

9. The aqueous pharmaceutical composition according to claim 7, characterized in that: The concentration of trehalose dihydrate was 80 mg / mL.

10. The aqueous pharmaceutical composition according to claim 7, characterized in that: The concentration of sodium acetate trihydrate is 1.7 mg / mL to 1.8 mg / mL.

11. The aqueous pharmaceutical composition according to claim 10, characterized in that: The concentration of sodium acetate trihydrate is 1.742 mg / mL.

12. The aqueous pharmaceutical composition according to claim 7, characterized in that: Add acetic acid to a pH of 5.0-5.

5.

13. An aqueous pharmaceutical composition for an anti-PD-1 antibody, comprising the following: (a) Prolgolimab at a concentration of 15 mg / mL to 40 mg / mL as an antibody; (b) Trehalose dihydrate at a concentration of 95 mg / mL to 105 mg / mL; (c) L-histidine at a concentration of 0.7–1.0 mg / mL; and (d) L-histidine hydrochloride at a concentration of 2.8–3.3 mg / mL.

14. The aqueous pharmaceutical composition according to claim 13, characterized in that: The concentration of Prolgolimab is 15 mg / mL to 25 mg / mL.

15. The aqueous pharmaceutical composition according to claim 14, characterized in that: The concentration of Prolgolimab was 20 mg / mL.

16. The aqueous pharmaceutical composition according to claim 13, characterized in that: The concentration of L-histidine was 0.92 mg / mL.

17. The aqueous pharmaceutical composition according to claim 13, characterized in that: The concentration of L-histidine hydrochloride was 2.96 mg / mL.

18. The aqueous pharmaceutical composition according to claim 13, characterized in that: The pH value of the composition is 5.5 to 6.

5.

19. The aqueous pharmaceutical composition according to claim 18, characterized in that: The pH value of the composition is 5.5 to 6.

0.

20. An aqueous pharmaceutical composition of anti-PD-1 antibody, comprising the following: (a) Prolgolimab at a concentration of 20 mg / mL as an antibody; (b) Trehalose dihydrate at a concentration of 100 mg / mL; (c) Sodium acetate trihydrate at a concentration of 1.7 mg / mL to 1.8 mg / mL; and (d) Add acetic acid to a pH of 5.

0.

21. The aqueous pharmaceutical composition for the anti-PD-1 antibody according to claim 20, comprising the following: (a) Prolgolimab at a concentration of 20 mg / mL as an antibody; (b) Trehalose dihydrate at a concentration of 100 mg / mL; (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL; and (d) Add acetic acid to a pH of 5.

0.

22. An aqueous pharmaceutical composition for an anti-PD-1 antibody, comprising the following: (a) Prolgolimab at a concentration of 20 mg / mL as an antibody; (b) Trehalose dihydrate at a concentration of 100 mg / mL; (c) L-histidine at a concentration of 0.7–1.0 mg / mL; and (d) L-histidine hydrochloride at a concentration of 2.8–3.3 mg / mL; (e) and the pH value of the composition is 5.5 to 6.

5.

23. The aqueous pharmaceutical composition for the anti-PD-1 antibody according to claim 22, comprising the following: (a) Prolgolimab at a concentration of 20 mg / mL as an antibody; (b) Trehalose dihydrate at a concentration of 100 mg / mL; (c) L-histidine at a concentration of 0.92 mg / mL; as well as (d) L-histidine hydrochloride at a concentration of 2.96 mg / mL; (e) and the pH value of the composition is 5.

5.

24. An aqueous pharmaceutical composition for an anti-PD-1 antibody, comprising the following: (a) Prolgolimab at a concentration of 20 mg / mL as an antibody; (b) Trehalose dihydrate at a concentration of 100 mg / mL; (c) Sodium acetate trihydrate at a concentration of 1.742 mg / mL; (d) Add acetic acid to a pH of 5.0; (e) Add water for injection to 1 mL.

25. The aqueous pharmaceutical composition according to any one of claims 1 to 24, characterized in that: The composition is to be administered parenterally.

26. The aqueous pharmaceutical composition according to claim 25, characterized in that: The composition is administered intravenously, subcutaneously, or intramuscularly.

27. The aqueous pharmaceutical composition according to any one of claims 1 to 24, characterized in that: The composition is contained in a small vial.

28. The aqueous pharmaceutical composition according to claim 27, characterized in that: The small medicine bottle is a glass medicine bottle.

29. The aqueous pharmaceutical composition according to claim 27, characterized in that: The volume of the small medicine bottle is 1mL to 50mL.

30. The aqueous pharmaceutical composition according to claim 27, characterized in that: The volume of the small medicine bottle is 5mL, 10mL, 15mL or 20mL.

31. The aqueous pharmaceutical composition according to any one of claims 1 to 24, characterized in that: The composition is placed inside a syringe.

32. The aqueous pharmaceutical composition according to claim 31, characterized in that: The syringe has a volume of 1 mL.

33. The aqueous pharmaceutical composition according to claim 31, characterized in that: The syringe has a volume of 2 mL.

34. The aqueous pharmaceutical composition according to any one of claims 1 to 24, characterized in that: The composition is placed in a pre-filled syringe.

35. The aqueous pharmaceutical composition according to claim 34, characterized in that: The pre-filled syringe has a volume of 1 mL.

36. The aqueous pharmaceutical composition according to claim 34, characterized in that: The pre-filled syringe has a volume of 2 mL.

37. Use of the aqueous pharmaceutical composition according to any one of claims 1 to 36 in the preparation of a medicament for treating malignant tumors in patients with corresponding needs, wherein the malignant tumor is selected from: inoperable melanoma or metastatic melanoma, inoperable non-small cell lung cancer or metastatic non-small cell lung cancer; non-squamous non-small cell lung cancer.

38. The use according to claim 37, wherein the aqueous pharmaceutical composition is administered at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

39. The use according to claim 37, wherein the aqueous pharmaceutical composition is administered at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

40. The use according to claim 37, wherein the aqueous pharmaceutical composition is applied every two weeks.

41. The use according to claim 37, wherein the aqueous pharmaceutical composition is applied every three weeks.

42. The use according to claim 37, wherein the aqueous pharmaceutical composition is administered every two weeks at a dose of 1 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

43. The use according to claim 37, wherein the aqueous pharmaceutical composition is administered every three weeks at a dose of 3 mg / kg body weight of the anti-PD-1 antibody Prolgolimab.

44. The use according to claim 37, wherein the aqueous pharmaceutical composition is administered parenterally.

45. The use according to claim 44, wherein the parenteral administration is intravenous, subcutaneous or intramuscular.

46. ​​The use according to claim 44, wherein the aqueous pharmaceutical composition is administered intravenously as an infusion.

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