Pharmaceutical compositions of humanized anti-CD40 antibodies

Abstract

The present disclosure relates to pharmaceutical compositions of anti-CD40 antibodies, such as humanized anti-CD40 antibody KPL-404. The pharmaceutical compositions include polysorbate 20, acetate, and a polar excipient. The pharmaceutical compositions can be used in various therapeutic, prophylactic, and diagnostic methods. The antibodies block the ability of CD40 to bind to CD154 without activating cells expressing CD40 (e.g., B cells).

Description

[Technical field]

[0001] (Sequence Description) This application contains a sequence listing filed electronically in Extensible Markup Language (XML) format, which is incorporated herein by reference in its entirety. Said XML copy, created on October 31, 2022, is named 51383-007WO2_Sequence_Listing_10_31_22.XML and is 20,931 bytes in size. [Background technology]

[0002] Suppression of the immune system, particularly the humoral immune system, is beneficial in organ transplantation and the treatment of autoimmune disorders.

[0003] One target for suppressing the immune system is the CD40 / CD154 interaction. CD40 is expressed mainly on the surface of B lymphocytes and other antigen-presenting cells (APCs) such as dendritic cells and macrophages. CD154 is expressed mainly on the surface of T cells. The interaction between these two proteins is associated with B cell activation, which induces cytokine expression and the expression of cell surface markers including CD23, CD80 and CD86. Antibodies (e.g., humanized antibodies) that target the CD40 / CD154 interaction have been developed. However, these antibodies or their antigen-binding fragments need to be formulated into pharmaceutical compositions suitable for in vivo administration for effective treatment. Summary of the Invention

[0004] In general, the present invention features a pharmaceutical composition containing a humanized anti-CD40 antibody or antigen-binding fragment thereof having a heavy chain variable region having an amino acid sequence having at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region having an amino acid sequence having at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to the amino acid sequence set forth in SEQ ID NO:10. In one embodiment, the humanized anti-CD40 antibody is KPL-404. The pharmaceutical composition includes polysorbate 20, acetate, and one or more polar excipients. The polar excipient may be or include, for example, a sugar, a polyol, or an amino acid. In some embodiments, the sugar is, for example, sucrose, trehalose, fructose, lactose, dextrose, or mannitol. In some embodiments, the polyol is, for example, polyethylene glycol or sorbitol. In some embodiments, the amino acid is one or more of alanine, arginine, aspartic acid, asparagine, carnitine, citrulline, ornithine, glycine, glutamic acid, glutamine, glycine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tyrosine, and valine. In some embodiments, the pharmaceutical composition comprises about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 45 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM) acetate. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate.

[0005] In some embodiments, the pharmaceutical composition comprises about 1 mM to about 200 mM (e.g., about 50 mM to about 150 mM, e.g., about 75 mM to about 125 mM, e.g., about 90 mM to about 110 mM, e.g., about 95 mM to about 105 mM, e.g., about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM) of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 50 mM of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 100 mM of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 150 mM of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 200 mM of a polar excipient.

[0006] In some embodiments, the pharmaceutical composition comprises about 1% (w / v) to about 10% (w / v) (e.g., about 2% (w / v) to about 8% (w / v), e.g., about 1% (w / v), 1.5% (w / v), 2% (w / v), 2.5% (w / v), 3% (w / v), 3.5% (w / v), 4% (w / v), 4.5% (w / v), 5% (w / v), 5.5% (w / v), 6% (w / v), 6.5% (w / v), 7% (w / v), 7.5% (w / v), 8% (w / v), 8.5% (w / v), 9% (w / v), 9.5% (w / v), 10% (w / v)) of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 2.5% (w / v) polar excipient. In some embodiments, the pharmaceutical composition comprises about 7% (w / v) polar excipient.

[0007] In some embodiments, the pharmaceutical composition comprises about 0.001% to about 0.1% (e.g., about 0.005% to 0.05%, e.g., about 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%) polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.02% polysorbate 20.

[0008] The composition may have a pH of about 5.0 to about 8.0, such as about 5.4 to about 6.5, such as about 5.4 to about 5.75, such as 5.4 to about 5.6, such as 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0, such as about 5.4.

[0009] In a first aspect, the present invention features a pharmaceutical composition comprising a humanized anti-CD40 antibody or antigen-binding fragment thereof, the antibody having a heavy chain variable region having an amino acid sequence having at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region having an amino acid sequence having at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to the amino acid sequence set forth in SEQ ID NO:10. In one embodiment, the humanized anti-CD40 antibody is KPL-404. The pharmaceutical composition includes polysorbate 20, acetate, and a polar excipient. For example, the polar excipient may include arginine and glutamate.

[0010] In some embodiments, the pharmaceutical composition comprises about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 45 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM) acetate. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate.

[0011] In some embodiments, the pharmaceutical composition comprises about 1 mM to about 200 mM (e.g., about 50 mM to about 150 mM, e.g., about 75 mM to about 125 mM, e.g., about 90 mM to about 110 mM, e.g., about 95 mM to about 105 mM, e.g., about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM) arginine. In some embodiments, the pharmaceutical composition comprises about 50 mM arginine. In some embodiments, the pharmaceutical composition comprises about 100 mM arginine.

[0012] In some embodiments, the pharmaceutical composition comprises about 1 mM to about 200 mM (e.g., about 50 mM to about 150 mM, e.g., about 50 mM to about 100 mM, e.g., about 75 mM to about 125 mM, e.g., about 90 mM to about 110 mM, e.g., about 95 mM to about 105 mM, e.g., about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM) of glutamate. In some embodiments, the pharmaceutical composition comprises about 50 mM glutamate. In some embodiments, the pharmaceutical composition comprises about 100 mM glutamate.

[0013] In some embodiments, the pharmaceutical composition comprises about 0.001% to about 0.1% (e.g., about 0.005% to 0.05%, e.g., about 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%) polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.02% polysorbate 20.

[0014] In some embodiments, the pharmaceutical composition comprises about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, e.g., about 50 mM acetate, and about 1 mM to about 200 mM (e.g., about 50 mM to about 150 mM, e.g., about 75 mM to about 125 mM, for example, about 90 mM to about 110 mM, for example, about 95 mM to about 105 mM, for example, about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM, for example, about 100 mM) arginine; About 1 mM to about 200 mM (for example, about 50 mM to about 150 mM, for example, about 75 mM to about 125 mM, for example, about 90 mM to about 110 mM, for example, about 95 mM to about 105 mM, for example, about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 1 90 mM, or 200 mM, e.g., about 100 mM) glutamate, and about 0.001% to about 0.1% (e.g., 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%, e.g., about 0.01%) polysorbate 20.The composition may have a pH of about 5.0 to about 8.0, such as about 5.4 to about 6.5, such as about 5.4 to about 5.75, such as 5.4 to about 5.6, such as 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0, such as about 5.4.

[0015] In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 100 mM arginine, 100 mM glutamate, and about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 100 mM arginine, 100 mM glutamate, and about 0.02% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 50 mM arginine, 50 mM glutamate, and about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 50 mM arginine, 50 mM glutamate, and about 0.02% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 100 mM arginine, 50 mM glutamate, and about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 100 mM arginine, 50 mM glutamate, and about 0.02% polysorbate 20. The composition may have a pH of about 5.4.

[0016] In a second aspect, the present invention features a pharmaceutical composition that includes a humanized anti-CD40 antibody or antigen-binding fragment thereof, having a heavy chain variable region having an amino acid sequence that has at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region having an amino acid sequence that has at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to the amino acid sequence set forth in SEQ ID NO:10. The pharmaceutical composition includes polysorbate 20, acetate, and a polar excipient. The polar excipient may be or include sucrose.

[0017] In some embodiments, the pharmaceutical composition comprises about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM) acetate. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate.

[0018] In some embodiments, the pharmaceutical composition comprises about 1% (w / v) to about 10% (w / v), e.g., 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, or 10% (w / v) sucrose. In some embodiments, the pharmaceutical composition comprises about 7% (w / v) sucrose.

[0019] In some embodiments, the pharmaceutical composition comprises about 0.001% to about 0.1% (e.g., about 0.005% to 0.05%, e.g., about 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%) polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.02% polysorbate 20.

[0020] In some embodiments, the pharmaceutical composition comprises about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, e.g., about 50 mM) acetate and about 1% (w / v) to about 10% (w / v), e.g., 1%, 2%, 3%, 4%, 5%, and about 0.001% to about 0.1% (e.g., 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%, e.g., about 0.01% or about 0.02%) polysorbate 20. The composition may have a pH of about 5.0 to about 8.0, such as about 5.4 to about 6.5, such as about 5.4 to about 5.75, such as 5.4 to about 5.6, such as 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0, such as about 5.4.

[0021] In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 7% (w / v) sucrose, and about 0.01% polysorbate 20. The composition may have a pH of about 5.4.

[0022] In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 7% (w / v) sucrose, and about 0.02% polysorbate 20. The composition may have a pH of about 5.4.

[0023] In a third aspect, the present invention features a pharmaceutical composition comprising a humanized anti-CD40 antibody or antigen-binding fragment thereof, having a heavy chain variable region having an amino acid sequence having at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to the amino acid sequence set forth in SEQ ID NO:9, and a light chain variable region having an amino acid sequence having at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to the amino acid sequence set forth in SEQ ID NO:10. The pharmaceutical composition comprises polysorbate 20, acetate, and a polar excipient. The polar excipient may be or may comprise sorbitol. The pharmaceutical composition may further comprise sodium chloride.

[0024] In some embodiments, the pharmaceutical composition comprises about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM) acetate. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate.

[0025] In some embodiments, the pharmaceutical composition comprises about 0.1% (w / v) sorbitol to about 5% (w / v), e.g., about 1 mM to about 5 mM, e.g., about 2 mM to about 3 mM, e.g., 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 2%, 3%, 4%, or 5% (w / v) sorbitol. In some embodiments, the pharmaceutical composition comprises about 2.5% (w / v) sorbitol.

[0026] In some embodiments, the pharmaceutical composition comprises about 1 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 50 mM to about 90 mM, e.g., about 60 mM to about 80 mM, e.g., about 65 mM to about 75 mM, e.g., about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM) sodium chloride. In some embodiments, the pharmaceutical composition comprises about 70 mM sodium chloride.

[0027] In some embodiments, the pharmaceutical composition comprises about 0.001% to about 0.1% (e.g., about 0.005% to 0.05%, e.g., about 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%) polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.02% polysorbate 20.

[0028] In some embodiments, the pharmaceutical composition comprises about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, e.g., about 50 mM acetate) and about 0.1% (w / v) sorbitol to about 5% (w / v), e.g., 0.2%, 0.3%, or 10% (w / v) sorbitol. %, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 2%, 3%, 4%, or 5% (w / v), for example, about 2.5% sorbitol, and about 1 mM to about 100 mM (for example, about 10 mM to about 90 mM, for example, about 50 mM to about 90 mM, for example, about 60 mM to about 80 mM, for example, about 65 mM to about 75 mM, for example, about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, e.g., about 70 mM) sodium chloride; and about 0.001% to about 0.1% (e.g., 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%, e.g., about 0.01% or about 0.02%) polysorbate 20. The composition may have a pH of about 5.0 to about 8.0, for example, about 5.4 to about 6.5, for example, about 5.4 to about 5.75, for example, 5.4 to about 5.6, for example, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0, for example, about 5.4.

[0029] In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 2.5% (w / v) sorbitol, about 70 mM NaCl, and about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 2.5% (w / v) sorbitol, about 70 mM NaCl, and about 0.02% polysorbate 20. The composition may have a pH of about 5.4.

[0030] In some embodiments, the pharmaceutical composition of any one of the first, second, and third aspects has a pH of about 5.0 to about 8.0, such as about 5.4 to about 6.5, such as about 5.4 to about 5.75, such as 5.4 to about 5.6, such as 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, and 8.0. For example, the pH may be about 5.4.

[0031] In some embodiments of any one of the first, second, and third aspects, the anti-CD40 antibody or antigen-binding fragment thereof is at a concentration of about 0.01 mg / mL to about 300 mg / mL, e.g., about 100 mg / mL to about 250 mg / mL, about 150 mg / mL to about 250 mg / mL, or about 100 mg / mL to about 200 mg / mL (e.g., 0.02 mg / mL, 0.03 mg / mL, 0.04 mg / mL, 0.05 mg / mL, 0.06 mg / mL, 0.07 mg / mL, 0.08mg / mL, 0.09mg / mL, 0.1mg / mL, 0.2mg / mL, 0.3mg / mL, 0.4mg / mL, 0.5mg / mL, 0.6mg / mL, 0.7mg / mL, 0.8mg / mL, 0.9mg / mL, 1m g / mL, 2mg / mL, 3mg / mL, 4mg / mL, 5mg / mL, 6mg / mL, 7mg / mL, 8mg / mL, 9mg / mL, or 10mg / mL, 20mg / mL, 30mg / mL, 40mg / mL, 50mg / mL , 60mg / mL, 70mg / mL, 80mg / mL, 90mg / mL, 100mg / mL, 105mg / mL, 110mg / mL, 115mg / mL, 120mg / mL, 125mg / mL, 130mg / mL, 135mg / mL, 140mg / mL, 145mg / mL, 150mg / mL, 155mg / mL, 160mg / mL, 165mg / mL, 170mg / mL, 175mg / mL, 180mg / mL, 185mg / mL, 190mg / mL, 1 In some embodiments, the anti-CD40 antibody or antigen-binding fragment thereof is at a concentration of about 200 mg / mL.

[0032] In some embodiments of any one of the first, second, and third aspects, the composition is formulated in a volume of about 0.1 mL to about 2.0 mL (e.g., about 0.1 mL, 0.2 mL, 0.3 mL, 0.4 mL, 0.5 mL, 0.6 mL, 0.7 mL, 0.8 mL, 0.9 mL, 1.0 mL, 1.1 mL, 1.2 mL, 1.3 mL, 1.4 mL, 1.5 mL, 1.6 mL, 1.7 mL, 1.8 mL, 1.9 mL, or 2.0 mL, 2.5 mL, 3.0 mL, 3.5 mL, 4.0 mL, 4.5 mL, 5.5 mL, or 5.5 mL). In some embodiments, the composition is formulated in a volume of about 2.0 mL.

[0033] In some embodiments of any one of the first, second, and third aspects, the composition comprises a volume of about 0.1 mL to about 2.0 mL (e.g., about 0.1 mL, 0.2 mL, 0.3 mL, 0.4 mL, 0.5 mL, 0.6 mL, 0.7 mL, 0.8 mL, 0.9 mL, 1.0 mL, 1.1 mL, 1.2 mL, 1.3 mL, 1.4 mL, 1.5 mL, 1.6 mL, 1.7 mL, 1.8 mL, 1.9 mL, or 2.0 mL, 2.5 mL, 3.0 mL, 4.0 mL, 5.0 mL, 6.0 mL, 7.0 mL, 8.0 mL, 9.0 mL, 10.0 mL, 11.0 mL, 12.0 mL, 13.0 mL, 14.0 mL, 15.0 mL, 16.0 mL, 17.0 mL, 18.0 mL, 19.0 mL, 20.0 mL, 25.0 mL, 26.0 mL, 27.0 mL, 28.0 mL, 29.0 mL, 30.0 mL, 31.0 mL, 32.0 mL, 33.0 mL, 34.0 mL, 35.0 mL, 36.0 mL, 37.0 mL, 38.0 mL, 39.0 mL, 40.0 mL, 41.0 mL, 42.0 mL, 43.0 mL, 44.0 mL, 45.0 mL, 46.0 mL, 47.0 mL, 48.0 mL, 49.0 mL, 50.0 mL, 51.0 mL, 52.0 mL, 53.0 mL, 54.0 mL, 55.0 mL, 56.0 mL and the antibody or antigen-binding fragment thereof is formulated in a volume of about 50 mg / mL to about 300 mg / mL (e.g., 50 mg / mL, 55 mg / mL, 60 mg / mL, 65 mg / mL, 70 mg / mL, 75 mg / mL, 80 mg / mL, 85 mg / mL, 90 mg / mL, 95 mg / mL, 100 mg / mL, 105 mg / mL). L, 110mg / mL, 115mg / mL, 120mg / mL, 125mg / mL, 130mg / mL, 135mg / mL, 140mg / mL, 145mg / mL, 150mg / mL, 155mg / mL, 160 mg / mL, 165mg / mL, 170mg / mL, 175mg / mL, 180mg / mL, 185mg / mL, 190mg / mL, 195mg / mL, 200mg / mL, 205mg / mL, 210mg / mL , 215mg / mL, 220mg / mL, 225mg / mL, 230mg / mL, 235mg / mL, 240mg / mL, 245mg / mL, 250mg / mL, 255mg / mL, 260mg / mL, 265mg / mL, 270mg / mL, 275mg / mL, 280mg / mL, 285mg / mL, 290mg / mL, 295mg / mL, or 300mg / mL, for example, in a volume of about 200mg / mL.

[0034] In some embodiments, at least 95% (e.g., at least 98%) of the humanized anti-CD40 antibody or antigen-binding fragment thereof in the composition is in monomeric form. In some embodiments, less than 5% (e.g., less than 2%) of the humanized anti-CD40 antibody or antigen-binding fragment thereof is present in the composition as a high molecular weight (HMW) species (e.g., in aggregated form).

[0035] In some embodiments, the amount of monomeric species of the humanized anti-CD40 antibody or antigen-binding fragment in the composition decreases to 95% or more (e.g., 98% or more) after 12 months of storage at 2-8° C. In some embodiments, the amount of HMW species of the humanized anti-CD40 antibody or antigen-binding fragment in the composition increases to 5% or less (e.g., 2% or less) after 12 months of storage at 2-8° C. In a fourth aspect, the invention features a kit including a pharmaceutical composition according to any one of the first, second, and third aspects described herein and instructions for use thereof.

[0036] A fifth aspect features the use of a pharmaceutical composition of any one of the first, second and third aspects described herein in the manufacture of a medicament for suppressing the immune system in a human subject.

[0037] A sixth aspect features a pharmaceutical composition of any one of the first, second and third aspects described herein for use in suppressing the immune system in a human subject.

[0038] In a seventh aspect, the invention features a method of suppressing the immune system of a human subject by administering to the subject (e.g., a subject in need thereof, such as a human subject) a pharmaceutical composition according to any one of the first, second, and third aspects described herein. [Brief description of the drawings]

[0039] The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Patent and Trademark Office upon request and payment of the necessary fee.

[0040] [Figure 1] FIG. 1 is a set of graphs showing the SEC results plotted for % major species, % HMW species, and % LMW species of KPL-404 for formulations A, 1, and 2. [Figure 2A]Figures 2A and 2B are graphs showing a comparison of particle concentrations from a biophysical characterization study: Figure 2A shows 200 mg / mL KPL-404 formulations A-C (no PS20 in formulation C). [Figure 2B] FIG. 2B shows high concentrations of KPL-404 in formulations AC. [Diagram 3] FIG. 3 is a graph showing viscosity versus shear rate of high concentration KPL-404 formulations. [Figure 4] FIG. 4 is a graph showing total KPL-404 species population by CG-MALS for Formulation A. [Diagram 5] FIG. 5 is a graph showing total KPL-404 species population by CG-MALS for Formulation B. [Figure 6] FIG. 6 is a graph showing total KPL-404 species population by CG-MALS for Formulation C. [Figure 7] FIG. 7 is a graph showing the exponential dependence of measured viscosity versus protein concentration for formulation A (circles), formulation B (squares), and formulation C (triangles). [Figure 8] FIG. 8 is a graph showing the Instron force profile, KPL-404 in Formulation A, with injection time from a standard Ypsomate autoinjector (2.25 mL). [Figure 9] FIG. 9 is a graph showing the Instron force profile, KPL-404 in Formulation B, with injection time from a standard Ypsomate autoinjector (2.25 mL). [Figure 10A] FIG. 10A is a graph showing the Instron force profile, KPL-404 in Formulation C, with injection time from a standard Ypsomate (FIG. 10A) autoinjector (2.25 mL). [Figure 10B] FIG. 10B is a graph showing the Instron force profile, KPL-404 in Formulation C, with injection time from a Ypsomate Pro (FIG. 10B) autoinjector (2.25 mL). [Figure 11]Figure 11 is a set of SEC chromatograms at t=0 (initial) for formulations A, B, and C. The right side shows an expanded view of the left panel. [Figure 12] Figure 12 is a set of SEC chromatograms of formulations A, B, and C at t=2w, 40° C. The right side shows an expanded view of the left panel. [Figure 13] FIG. 13 is a graph showing SEC percent major as a function of time for 5° C. storage. [Figure 14] FIG. 14 is a graph showing SEC percent major as a function of time upon storage at 25° C. [Figure 15] FIG. 15 is a graph showing SEC percent major as a function of time for 40° C. storage. [Figure 16] FIG. 16 is a graph showing SEC percent major as a function of time upon -70° C. storage. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0041] The present disclosure relates to pharmaceutical compositions containing anti-CD40 antibodies and antigen-binding fragments of the antibodies that can be used in various therapeutic, prophylactic, diagnostic and other methods. The pharmaceutical compositions contain antibodies or antigen-binding fragments thereof that can interfere with the ability of CD40 to bind to CD154 and can do so without activating cells (e.g., B cells) that express CD40. The pharmaceutical compositions described herein can be used to treat autoimmune diseases and disorders. Generally, the antibodies or antigen-binding fragments thereof are derived from the murine 2C10 antibody and its humanized variants, for example, as described in PTC publications WO2012 / 125569 and WO2017 / 040932, which are incorporated herein by reference in their entirety, including the sequences described therein. The pharmaceutical compositions contain combinations of particular carriers and excipients that impart surprisingly beneficial properties to the compositions, including the ability to formulate the drug (anti-CD40 antibodies and antigen-binding fragments thereof) at high concentrations (as described herein), to increase the stability of the composition (e.g., extended shelf life), to reduce drug aggregation, and to improve the viscosity parameters of the composition. The components of the pharmaceutical compositions are described in more detail below.

[0042] Antibodies and Antibody-Binding Fragments The pharmaceutical compositions described herein include antibodies or antigen-binding fragments thereof derived from the murine 2C10 antibody, including, for example, but not limited to, the humanized anti-CD40 antibody identified as KPL-404. The heavy chain variable regions, light chain variable regions, and CDRs of certain anti-CD40 antibodies described herein are shown in Table 1. [Table 1-1] [Table 1-2] [Table 1-3]

[0043] In certain embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising an amino acid sequence at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, about 70%, about 75%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% identical to the amino acid sequence of the heavy chain variable region set forth in SEQ ID NO: 9. In certain embodiments, the antibody comprises a heavy chain amino acid sequence set forth in SEQ ID NO:11.

[0044] In certain embodiments, the antibody or antigen-binding fragment thereof comprises a light chain variable region comprising an amino acid sequence at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, about 70%, about 75%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% identical to the amino acid sequence of the light chain variable region set forth in SEQ ID NO: 10. In certain embodiments, the antibody comprises a light chain amino acid sequence set forth in SEQ ID NO:12.

[0045] In certain embodiments, the antibodies or antigen-binding fragments thereof each have an amino acid sequence at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, about 70%, about 75%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% identical to the amino acid sequence of the heavy chain variable region set forth in SEQ ID NO:9. and a light chain variable region comprising an amino acid sequence at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, about 70%, about 75%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% identical to the variable light chain amino acid sequence set forth in SEQ ID NO: 10. In certain embodiments, the antibody comprises a heavy chain amino acid sequence set forth in SEQ ID NO: 11 and a light chain amino acid sequence set forth in SEQ ID NO: 12. In certain embodiments, the anti-CD40 antibody is KPL-404.

[0046] In certain embodiments, the antibody or antigen-binding fragment thereof used to generate the pharmaceutical compositions described herein has a sequence identity that is at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, about 70%, about 75%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or more identical to the amino acid sequence of the heavy chain variable region set forth in any one of SEQ ID NOs: 13, 14, 16, 17 or 18. and a light chain variable region comprising an amino acid sequence having at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, about 70%, about 75%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% sequence identity to the variable light chain amino acid sequence set forth in any one of SEQ ID NOs: 15, 19, 20 or 21.

[0047] In certain embodiments, the heavy chain variable region of the antibody or antigen-binding fragment thereof comprises complementarity determining regions (CDRs) that are at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, about 70%, about 75%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% identical to the CDRs of the heavy chain variable region of the KPL-404 antibody (CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 3, 4, 5, respectively).

[0048] In certain embodiments, the light chain variable region of the antibody or antigen-binding fragment thereof comprises CDRs that are at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, about 70%, about 75%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or about 100% identical to the CDRs of the light chain variable region of the KPL-404 antibody (CDR1, CDR2 and CDR3 set forth in SEQ ID NOs: 6, 7, 8, respectively).

[0049] In certain embodiments, the heavy chain comprises the CDRs set forth in SEQ ID NOs: 3-5, respectively, and the light chain comprises the CDRs set forth in SEQ ID NOs: 6-8, respectively.

[0050] In certain embodiments, the heavy chain has at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to SEQ ID NO:9 and the CDRs set forth in SEQ ID NOs:3-5, respectively, and the light chain has at least 80% (e.g., at least 85%, 90%, 95%, 97%, 99%, or 100%) sequence identity to SEQ ID NO:10 and the CDRs set forth in SEQ ID NOs:6-8, respectively.

[0051] Also within the scope of the present disclosure are antibodies or antigen-binding fragments in which specific amino acids are substituted, deleted, or added. These changes have no substantial effect on the biological properties of the peptide, such as binding activity. For example, an antibody may have amino acid substitutions in the framework region, such as to improve binding to an antigen. In another example, a selected small number of acceptor framework residues may be replaced by the corresponding donor amino acids. The donor framework may be a mature or germline human antibody framework sequence or a consensus sequence. Guidance on how to make phenotypically silent amino acid substitutions can be found, for example, in Bowie et al. (Science, 247: 1306-1310, 1990), Cunningham et al. (Science, 244: 1081-1085, 1989), Ausubel (ed.) (Current Protocols in Molecular Biology, John Wiley and Sons, Inc., 1994), T. Maniatis, E. F. Fritsch and J. Sambrook (Molecular Cloning: A Laboratory Manual, Cold Spring Harbor laboratory, Cold Spring Harbor, NY, 1989), Pearson (Methods Mol. Biol. 243:307-31, 1994), and Gonnet et al. (Science 256:1443-45, 1992), each of which is incorporated herein by reference.

[0052] The peptides described herein may be functionally active variants of the antibodies or antigen-binding fragments thereof disclosed herein, e.g., in which less than about 30%, about 25%, about 20%, about 15%, about 10%, about 5%, or about 1% of the amino acid residues have been substituted or deleted, but which retain essentially the same immunological properties, including but not limited to binding to CD40.

[0053] Antibodies or antigen-binding fragments thereof may also include variants, analogs, orthologs, homologs, and derivatives of the peptides that exhibit biological activity, such as binding an antigen, such as CD40. The peptides may contain one or more homologs of an amino acid (including, for example, unnatural amino acids, amino acids that occur naturally only in unrelated biological systems, modified amino acids from mammalian systems, etc.), peptides with substituted bonds, as well as other modifications known in the art.

[0054] The antibody or antigen-binding fragment thereof may be derivatized or conjugated to another functional molecule. For example, the antibody may be functionally conjugated (by chemical coupling, genetic fusion, non-covalent interactions) to one or more other molecular entities, such as another antibody, a detectable agent, an immunosuppressant, a cytotoxic agent, a pharmaceutical agent, a protein or peptide that can mediate association with other molecules (such as the core region of streptavidin or polyhistidine tabs), an amino acid linker, a signal sequence, an immunogenic carrier, or a ligand useful for protein purification (such as glutathione-S-transferase, histidine tags, and Staphylococcus protein A). Cytotoxic agents may include radioisotopes, chemotherapeutic agents, and toxins and fragments thereof, such as enzymatically active toxins of bacterial, fungal, plant, or animal origin. Such cytotoxic agents may be conjugated to the humanized antibodies of the present disclosure using standard procedures and used, for example, to treat patients who are indicated for therapy with the antibody.

[0055] One type of derivatized protein is produced by crosslinking two or more proteins (of the same or different types). Suitable crosslinkers include those that are heterobifunctional, having two distinct reactive groups separated by a suitable spacer (e.g., m-maleimidobenzoyl-N-hydroxysuccinimide ester), or homobifunctional (e.g., disuccinimidyl suberate) crosslinkers. Useful detectable agents with which proteins can be derivatized (or labeled) include fluorescent agents, various enzymes, prosthetic groups, luminescent agents, bioluminescent agents, and radioactive agents. Non-limiting exemplary fluorescent detectable agents include fluorescein, fluorescein isothiocyanate, rhodamine, and phycoerythrin. Proteins or antibodies can also be derivatized with detectable enzymes, such as alkaline phosphatase, horseradish peroxidase, beta-galactosidase, acetylcholinesterase, glucose oxidase, and the like. Proteins also can be derivatized with prosthetic groups (eg, streptavidin / biotin and avidin / biotin).

[0056] In another embodiment, the humanized anti-CD40 antibody or fragment thereof is used unlabeled and detected with a labeled antibody that binds to the humanized anti-CD40 antibody or fragment thereof. Pharmaceutical Compositions

[0057] The disclosure features a pharmaceutical composition containing an antibody or antigen-binding fragment thereof described herein (e.g., an antibody or antigen-binding fragment thereof containing a heavy chain variable region having at least 80%, 85%, 90%, 95%, 97%, 99%, or 100% sequence identity to SEQ ID NO:9 and a light chain variable region having at least 80%, 85%, 90%, 95%, 97%, 99%, or 100% sequence identity to SEQ ID NO:10, formulated with a pharma- ceutically acceptable carrier). The disclosure also features a pharmaceutical composition containing an antibody or antigen-binding fragment thereof described herein (e.g., an antibody or antigen-binding fragment thereof containing a heavy chain having at least 80%, 85%, 90%, 95%, 97%, 99%, or 100% sequence identity to SEQ ID NO: 11 and a light chain having at least 80%, 85%, 90%, 95%, 97%, 99%, or 100% sequence identity to SEQ ID NO: 12, formulated with a pharma- ceutically acceptable carrier). The pharmaceutical composition containing the antibody or antigen-binding fragment thereof can be used for the treatment of a disease or disorder, e.g., as described herein. The pharmaceutical composition includes polysorbate 20, acetate, and one or more polar excipients. The polar excipient may be or include, e.g., a sugar, a polyol, or an amino acid. In some embodiments, the sugar is, e.g., sucrose, trehalose, fructose, lactose, dextrose, or mannitol. In some embodiments, the polyol is, for example, polyethylene glycol or sorbitol. In some embodiments, the amino acid is one or more of alanine, arginine, aspartic acid, asparagine, carnitine, citrulline, ornithine, glycine, glutamic acid, glutamine, glycine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tyrosine, and valine. The acetate may include, for example, sodium acetate (e.g., in salt form) or acetate in its ionic form. In some embodiments, the polar excipient is, for example, sucrose, arginine, glutamate, sorbitol, or a combination thereof. In some embodiments, the polar excipient is sucrose.In some embodiments, the polar excipient is arginine. In some embodiments, the polar excipient is glutamate. In some embodiments, the polar excipient is a mixture of arginine and glutamate. In some embodiments, the polar excipient is sorbitol.

[0058] In some embodiments, the pharmaceutical composition comprises about 5 mM to about 100 (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 45 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM) acetate. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate.

[0059] In some embodiments, the pharmaceutical composition comprises about 1 mM to about 200 mM (e.g., about 50 mM to about 150 mM, e.g., about 75 mM to about 125 mM, e.g., about 90 mM to about 110 mM, e.g., about 95 mM to about 105 mM, e.g., about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM) of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 50 mM of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 100 mM of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 150 mM of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 200 mM of a polar excipient.

[0060] In some embodiments, the pharmaceutical composition comprises about 1% (w / v) to about 10% (w / v) (e.g., about 2% (w / v) to about 8% (w / v), e.g., about 1% (w / v), 1.5% (w / v), 2% (w / v), 2.5% (w / v), 3% (w / v), 3.5% (w / v), 4% (w / v), 4.5% (w / v), 5% (w / v), 5.5% (w / v), 6% (w / v), 6.5% (w / v), 7% (w / v), 7.5% (w / v), 8% (w / v), 8.5% (w / v), 9% (w / v), 9.5% (w / v), 10% (w / v)) of a polar excipient. In some embodiments, the pharmaceutical composition comprises about 2.5% (w / v) polar excipient. In some embodiments, the pharmaceutical composition comprises about 7% (w / v) polar excipient.

[0061] In some embodiments, the pharmaceutical composition comprises about 0.001% to about 0.1% (e.g., about 0.005% to 0.05%, e.g., about 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%) polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.02% polysorbate 20.

[0062] The composition may have a pH of about 5.0 to about 8.0, such as about 5.4 to about 6.5, such as about 5.4 to about 5.75, such as 5.4 to about 5.6, such as 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0, such as about 5.4.

[0063] The pharmaceutical composition may be at about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, For example, about 50 mM) acetate, and about 1 mM to about 200 mM (for example, about 50 mM to about 150 mM, for example, about 75 mM to about 125 mM, for example, about 90 mM to about 110 mM, for example, about 95 mM to about 105 mM, for example, about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 6 0 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM, e.g., about 100 mM) of a polar excipient and about 0.001% to about 0.1% (e.g., 0.001%, about 0.002%, 0.003% , 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%, e.g., about 0.01% or about 0.02%) of polysorbate 20. The pharmaceutical composition may have a pH of about 5.0 to about 8.0, for example, about 5.4 to about 6.5, for example, about 5.4 to about 5.75, for example, 5.4 to about 5.6, for example, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0. For example, the pH may be about 5.4.

[0064] The pharmaceutical composition may, for example, contain an amount of an anti-CD40 antibody or antigen-binding fragment thereof described herein (e.g., KPL-404), for example, about 100 mg / mL to about 300 mg / mL, for example, about 100 mg / mL to about 250 mg / mL, about 150 mg / mL to about 250 mg / mL, or about 100 mg / mL to about 200 mg / mL (e.g., about 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, 150 mg / mL, 155 mg / mL, 160 mg / mL, L, 165 mg / mL, 170 mg / mL, 175 mg / mL, 180 mg / mL, 185 mg / mL, 190 mg / mL, 195 mg / mL, 200 mg / mL, 205 mg / mL, 210 mg / mL, 215 mg / mL, 220 mg / mL, 225 mg / mL, 230 mg / mL, 235 mg / mL, 240 mg / mL, 245 mg / mL, and 250 mg / mL, 255 mg / mL, 260 mg / mL, 265 mg / mL, 270 mg / mL, 275 mg / mL, 280 mg / mL, 285 mg / mL, 290 mg / mL, 295 mg / mL, or 300 mg / mL).

[0065] In some embodiments, at least 95% (e.g., at least 98%) of the humanized anti-CD40 antibody or antigen-binding fragment thereof in the composition is in monomeric form. In some embodiments, less than 5% (e.g., less than 2%) of the humanized anti-CD40 antibody or antigen-binding fragment thereof is present in the composition as a high molecular weight (HMW) species (e.g., in aggregated form).

[0066] In some embodiments, the amount of monomeric species of the humanized anti-CD40 antibody or antigen-binding fragment in the composition decreases to 95% or more (e.g., 98% or more) after 12 months of storage at 2-8° C. In some embodiments, the amount of HMW species of the humanized anti-CD40 antibody or antigen-binding fragment in the composition increases to 5% or less (e.g., 2% or less) after 12 months of storage at 2-8° C.

[0067] Formulation C Formulation C: 200 mg / mL KPL-404 in 50 mM acetate, 100 mM arginine, 100 mM glutamate, 0.02% polysorbate 20, pH 5.4.

[0068] Pharmaceutical compositions of the present disclosure may also include modifications to Formulation C. For example, the pharmaceutical composition can include different amounts of polysorbate 20, acetate, arginine, and glutamate than Formulation C. For example, the pharmaceutical composition can include about 5 mM to about 100 mM (e.g., about 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM) acetate, such as about 50 mM acetate. The pharmaceutical composition can include about 1 mM to about 200 mM arginine (e.g., about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM), such as about 50 mM or about 100 mM arginine. The pharmaceutical composition can include about 1 mM to about 200 mM (e.g., about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM) glutamate, such as about 50 mM or about 100 mM glutamate.

[0069] In some embodiments, the pharmaceutical composition comprises about 0.001% to about 0.1% polysorbate 20 (e.g., about 0.005% to 0.05%, e.g., about 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%), such as about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 0.02% polysorbate 20.

[0070] The pharmaceutical composition may comprise about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, for example, about 25 mM to about 75 mM, for example, about 40 mM to about 60 mM, for example, about 45 mM to about 55 mM, for example, about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, for example, about 50 mM) of acetate and about 1 mM to about 200 mM (e.g., about 50 mM to about 150 mM, for example, about 75 mM to about 125 mM) of riboflavin. M, for example, about 90 mM to about 110 mM, for example, about 95 mM to about 105 mM, for example, about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM, for example, about 100 mM) arginine and about 1 mM to about 200 mM (e.g. For example, about 50 mM to about 150 mM, for example, about 75 mM to about 125 mM, for example, about 90 mM to about 110 mM, for example, about 95 mM to about 105 mM, for example, about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM, For example, about 100 mM) glutamate and about 0.001% to about 0.1% (e.g., 0.001%, about 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%, e.g., about 0.01% or about 0.02%) polysorbate 20.

[0071] The pharmaceutical composition can include about 50 mM acetate, about 100 mM arginine, about 100 mM glutamate, and about 0.01% polysorbate 20. The pharmaceutical composition can include about 50 mM acetate, about 100 mM arginine, about 100 mM glutamate, and about 0.02% polysorbate 20. In some embodiments, the pharmaceutical composition includes about 50 mM acetate, about 50 mM arginine, 50 mM glutamate, and about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition includes about 50 mM acetate, about 50 mM arginine, 50 mM glutamate, and about 0.02% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 100 mM arginine, 50 mM glutamate, and about 0.01% polysorbate 20. In some embodiments, the pharmaceutical composition comprises about 50 mM acetate, about 100 mM arginine, 50 mM glutamate, and about 0.02% polysorbate 20. The pharmaceutical composition may have a pH of about 5.0 to about 8.0, for example, about 5.4 to about 6.5, for example, about 5.4 to about 5.75, for example, 5.4 to about 5.6, for example, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0. For example, the pH may be about 5.4.

[0072] The pharmaceutical composition may, for example, contain an amount of an anti-CD40 antibody or antigen-binding fragment thereof described herein (e.g., KPL-404), for example, about 100 mg / mL to about 300 mg / mL, for example, about 100 mg / mL to about 250 mg / mL, about 150 mg / mL to about 250 mg / mL, or about 100 mg / mL to about 200 mg / mL (e.g., about 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, 150 mg / mL, 155 mg / mL, 160 mg / mL, L, 165 mg / mL, 170 mg / mL, 175 mg / mL, 180 mg / mL, 185 mg / mL, 190 mg / mL, 195 mg / mL, 200 mg / mL, 205 mg / mL, 210 mg / mL, 215 mg / mL, 220 mg / mL, 225 mg / mL, 230 mg / mL, 235 mg / mL, 240 mg / mL, 245 mg / mL, and 250 mg / mL, 255 mg / mL, 260 mg / mL, 265 mg / mL, 270 mg / mL, 275 mg / mL, 280 mg / mL, 285 mg / mL, 290 mg / mL, 295 mg / mL, or 300 mg / mL).

[0073] Formulation A Formulation A: 200 mg / mL KPL-404 in 50 mM acetate, 7% (w / v) sucrose, 0.01% polysorbate 20, pH 5.4.

[0074] The pharmaceutical compositions of the present disclosure may also include modifications to Formulation A. For example, the pharmaceutical composition may include polysorbate 20, acetate, and sucrose. For example, the pharmaceutical composition may include about 5 mM to about 100 mM acetate (e.g., about 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM), such as about 50 mM acetate. The pharmaceutical composition may include about 1% (w / v) to about 10% (w / v), such as about 7% (w / v) sucrose, e.g., 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, or 10% (w / v). The pharmaceutical composition can include about 0.001% to about 0.1% (e.g., 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%) polysorbate 20, such as about 0.01% or about 0.02% polysorbate 20.

[0075] The pharmaceutical composition may comprise about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, e.g., about 50 mM) of acetate and about 1% (w / v) to about 10% (w / v), e.g., 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 82%, 83%, 84%, 8 %, 8%, 9%, or 10% (w / v), e.g., about 7% sucrose, and about 0.001% to about 0.1% (e.g., 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%, e.g., about 0.01%) polysorbate 20.

[0076] The pharmaceutical composition can include about 50 mM acetate, about 7% (w / v) sucrose, and about 0.01% polysorbate 20. The pharmaceutical composition can include about 50 mM acetate, about 7% (w / v) sucrose, and about 0.02% polysorbate 20.

[0077] The pharmaceutical composition may have a pH of about 5.0 to about 8.0, for example, about 5.4 to about 6.5, for example, about 5.4 to about 5.75, for example, 5.4 to about 5.6, for example, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0. For example, the pH may be about 5.4.

[0078] The pharmaceutical composition may, for example, contain an amount of an anti-CD40 antibody or antigen-binding fragment thereof described herein (e.g., KPL-404), for example, about 100 mg / mL to about 300 mg / mL, for example, about 100 mg / mL to about 250 mg / mL, about 150 mg / mL to about 250 mg / mL, or about 100 mg / mL to about 200 mg / mL (e.g., about 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, 150 mg / mL, 155 mg / mL, 160 mg / mL, L, 165 mg / mL, 170 mg / mL, 175 mg / mL, 180 mg / mL, 185 mg / mL, 190 mg / mL, 195 mg / mL, 200 mg / mL, 205 mg / mL, 210 mg / mL, 215 mg / mL, 220 mg / mL, 225 mg / mL, 230 mg / mL, 235 mg / mL, 240 mg / mL, 245 mg / mL, and 250 mg / mL, 255 mg / mL, 260 mg / mL, 265 mg / mL, 270 mg / mL, 275 mg / mL, 280 mg / mL, 285 mg / mL, 290 mg / mL, 295 mg / mL, or 300 mg / mL).

[0079] Formulation B Formulation B: 200 mg / mL KPL-404 in 50 mM acetate, 2.5% (w / v) sorbitol, 70 mM NaCl, 0.01% polysorbate 20, pH 5.4.

[0080] The pharmaceutical compositions of the present disclosure may also include modifications to Formulation B. For example, the pharmaceutical composition may include polysorbate 20, acetate, sorbitol, and sodium chloride. For example, the pharmaceutical composition may include about 5 mM to about 100 mM (e.g., about 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM) acetate, such as about 50 mM acetate. The pharmaceutical composition can include about 0.1% (w / v) sorbitol to about 5% (w / v), e.g., 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, or 5% (w / v) sorbitol, such as about 2.5% (w / v) sorbitol. The pharmaceutical composition can include about 1 mM to about 100 mM sodium chloride, such as about 70 mM sodium chloride (e.g., about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM). The pharmaceutical composition can include about 0.001% to about 0.1% (e.g., 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%) polysorbate 20, such as about 0.01% polysorbate 20 or about 0.02% polysorbate 20.

[0081] The pharmaceutical composition may contain about 5 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 25 mM to about 75 mM, e.g., about 40 mM to about 60 mM, e.g., about 45 mM to about 55 mM, e.g., about 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, e.g., about 50 mM) acetic acid. Salt, about 0.1% (w / v) sorbitol to about 5% (w / v), (e.g., 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 2%, 3%, 4%, or 5% (w / v), e.g., about 2.5%) sorbitol, and about 1 mM to about 100 mM (e.g., about 10 mM to about 90 mM, e.g., about 50 mM to about 90 mM) sorbitol. M, for example, about 60 mM to about 80 mM, for example, about 65 mM to about 75 mM, for example, about 5 mM, about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, or 100 mM, for example, about 70 mM) sodium chloride, and about 0.001% to about 0.1% (for example, 0. and 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, or 0.1%, e.g., about 0.01% or about 0.02%, of polysorbate 20.

[0082] The pharmaceutical composition can include about 50 mM acetate, about 2.5% (w / v) sorbitol, about 70 mM NaCl, and about 0.01% polysorbate 20. The pharmaceutical composition can include about 50 mM acetate, about 2.5% (w / v) sorbitol, about 70 mM NaCl, and about 0.02% polysorbate 20.

[0083] The pharmaceutical composition may have a pH of about 5.0 to about 8.0, for example, about 5.4 to about 6.5, for example, about 5.4 to about 5.75, for example, 5.4 to about 5.6, for example, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9 and 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 and 8.0. For example, the pH may be about 5.4.

[0084] The pharmaceutical composition may, for example, contain an amount of an anti-CD40 antibody or antigen-binding fragment thereof described herein (e.g., KPL-404), for example, about 100 mg / mL to about 300 mg / mL, for example, about 100 mg / mL to about 250 mg / mL, about 150 mg / mL to about 250 mg / mL, or about 100 mg / mL to about 200 mg / mL (e.g., about 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, 150 mg / mL, 155 mg / mL, 160 mg / mL, L, 165 mg / mL, 170 mg / mL, 175 mg / mL, 180 mg / mL, 185 mg / mL, 190 mg / mL, 195 mg / mL, 200 mg / mL, 205 mg / mL, 210 mg / mL, 215 mg / mL, 220 mg / mL, 225 mg / mL, 230 mg / mL, 235 mg / mL, 240 mg / mL, 245 mg / mL, and 250 mg / mL, 255 mg / mL, 260 mg / mL, 265 mg / mL, 270 mg / mL, 275 mg / mL, 280 mg / mL, 285 mg / mL, 290 mg / mL, 295 mg / mL, or 300 mg / mL).

[0085] Route of administration The compositions of the present disclosure can be formulated for administration by various methods known in the art. As will be appreciated by those skilled in the art, the route and / or mode of administration will vary depending on the desired results. Administration may be parenteral, intravenous, intrathecal, subcutaneous, oral, topical, local, intramuscular, intradermal, transdermal, subcutaneous, buccal, rectal, spinal, or epidermal. Intravenous delivery by continuous infusion is one exemplary method for administering the present antibody or antigen-binding fragment thereof.

[0086] In order to administer the agent by a certain administration route, it may be necessary to coat the agent with a material or to co-administer the agent with a material to prevent inactivation.For example, the agent may be administered to a subject with a suitable carrier, such as liposome, or diluent.Pharmaceutically acceptable diluents include physiological saline and aqueous buffer solutions.

[0087] Parenteral administration means modes of administration other than enteral and topical administration, usually by injection, and can include, but is not limited to, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injection and infusion.

[0088] Indications Pharmaceutical compositions containing the antibodies or antigen-binding fragments described herein have therapeutic, prophylactic, and / or diagnostic utility in vitro and in vivo. The antibodies or antigen-binding fragments thereof can be administered to a subject as part of an in vivo (e.g., therapeutic or prophylactic) protocol. For in vivo embodiments, the contacting step includes administering an anti-CD40 antibody or a portion thereof to a subject under conditions effective to allow binding of the antibody or antigen-binding fragment thereof to CD40 in the subject. The antibodies or antigen-binding fragments thereof can be administered to reduce the likelihood of transplant rejection or to prolong the period before transplant rejection, to induce immunosuppression, or to treat immune (e.g., autoimmune) disorders.

[0089] In one aspect, the pharmaceutical compositions described herein can be used to suppress the immune system of a human subject or can be used to manufacture a drug for suppressing the immune system of a human subject.The pharmaceutical compositions described herein can be used in any situation where immunosuppression is desired (e.g., transplant rejection or autoimmune disorders).These antibodies are particularly useful in the treatment of transplant rejection, for example, by reducing the likelihood that a particular transplant will be rejected by the host or by increasing the time until rejection occurs.

[0090] The pharmaceutical compositions described herein may also be used to treat autoimmune disorders, hi one embodiment, an autoimmune disorder may be associated with or caused by the presence of autoantibodies.

[0091] In another embodiment, the pharmaceutical compositions can be used to treat various disorders associated with the expression of CD40.

[0092] The disorder may be any condition that would benefit from treatment with the antibody or fragment thereof. This includes chronic and acute disorders or diseases, including pathological conditions that predispose the mammal to the disorder in question. Non-limiting examples of disorders that may be treated herein include autoimmune diseases, immune disorders, inflammatory disorders, and cancer.

[0093] Compositions and Kits The disclosure also features compositions and kits containing materials useful for the treatment of conditions or disorders described herein. The compositions or kits may include a container containing a pharmaceutical composition as described herein (e.g., one of formulations C, A, or B containing an amount of an anti-CD40 antibody or antigen-binding fragment thereof, such as about 100 mg / mL to about 300 mg / mL), and optionally a label. Suitable containers include, for example, bottles, vials, syringes, prefilled syringes, autoinjectors, and test tubes. The container may be formed from a variety of materials, such as glass or plastic. The container may hold a pharmaceutical composition as described herein, for example, effective for treating a condition, and may have a sterile access port. For example, the container may be an intravenous solution bag or vial having a stopper pierceable by a hypodermic needle. A label on or associated with the container indicates that the composition is used to treat a selected condition. The kit may further include a second container having a pharma- ceutically acceptable buffer, such as phosphate buffered saline, Ringer's solution, and dextrose solution. The kit may further include other materials desirable from a commercial and user standpoint, including other buffers, diluents, filters, needles, syringes, and package inserts with instructions for use.

[0094] The kit or composition can contain an antibody or antigen-binding fragment thereof as described herein formulated to about 100 mg / mL to about 300 mg / mL. In one embodiment, the composition is formulated to about 200 mg / mL of the antibody or antigen-binding fragment thereof. The antibody or antigen-binding fragment thereof may be formulated to a concentration of about 0.01 mg / mL to about 300 mg / mL (e.g., 0.02 mg / mL, 0.03 mg / mL, 0.04 mg / mL, 0.05 mg / mL, 0.06 mg / mL, 0.07 mg / mL, 0.08 mg / mL, 0.09 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.4 mg / mL, 0.5 mg / mL, 0. 6mg / mL, 0.7mg / mL, 0.8mg / mL, 0.9mg / mL, 1mg / mL, 2mg / mL, 3mg / mL, 4mg / mL, 5mg / mL, 6mg / mL, 7mg / mL, 8mg / mL, 9 mg / mL, or 10mg / mL, 20mg / mL, 30mg / mL, 40mg / mL, 50mg / mL, 60mg / mL, 70mg / mL, 80mg / mL, 90mg / mL, 100mg / mL, 10 5mg / mL, 110mg / mL, 115mg / mL, 120mg / mL, 125mg / mL, 130mg / mL, 135mg / mL, 140mg / mL, 145mg / mL, 150mg / mL, 155 mg / mL, 160mg / mL, 165mg / mL, 170mg / mL, 175mg / mL, 180mg / mL, 185mg / mL, 190mg / mL, 195mg / mL, 200mg / mL, 205mg The concentration may be 210 mg / mL, 215 mg / mL, 220 mg / mL, 225 mg / mL, 230 mg / mL, 235 mg / mL, 240 mg / mL, 245 mg / mL, 250 mg / mL, 255 mg / mL, 260 mg / mL, 265 mg / mL, 270 mg / mL, 275 mg / mL, 280 mg / mL, 285 mg / mL, 290 mg / mL, 295 mg / mL, or 300 mg / mL.

[0095] The composition may be formulated in a single-use vial having an extractable volume of about 0.5 mL to about 2.0 mL (e.g., 1.0 mL or 2.0 mL). The composition may also be formulated in a pre-filled syringe having an injectable volume of about 0.5 mL to about 2.0 mL (e.g., 1.0 mL or 2.0 mL). The composition may also be formulated in an autoinjector having an injectable volume of about 0.5 mL to about 2.0 mL (e.g., 1.0 mL or 2.0 mL).

[0096] The composition can be formulated in a volume of about 0.1 mL to about 10.0 mL (e.g., about 0.5 to about 5 mL, e.g., about 1.0 mL to about 2.0 mL, e.g., 0.1 mL, 0.2 mL, 0.3 mL, 0.4 mL, 0.5 mL, 0.6 mL, 0.7 mL, 0.8 mL, 0.9 mL, 1.0 mL, 1.1 mL, 1.2 mL, 1.3 mL, 1.4 mL, 1.5 mL, 1.6 mL, 1.7 mL, 1.8 mL, 1.9 mL, 2.0 mL, 3.0 mL, 4.0 mL, 5.0 mL, 6.0 mL, 7.0 mL, 8.0 mL, 9.0 mL, or 10.0 mL), such as a volume of about 2.0 mL.

[0097] Additional components that may be included in the kit are, for example, instructions for use, other reagents, therapeutic agents, or agents useful for labeling or coupling the antibody to a therapeutic agent or other materials for preparing the antibody for administration, a pharma- ceutically acceptable carrier, and a device or other materials for administration to a subject. EXAMPLES

[0098] The following examples of specific modes for carrying out the present disclosure are offered for illustrative purposes only and are not intended to limit the scope of the disclosure in any way. EXAMPLES

[0099] Described herein are studies that were performed to evaluate formulations that would allow for increasing the concentration of KPL-404 to at least 100 mg / mL.

[0100] Unless otherwise stated, the candidate formulations described herein include KPL-404 and the following excipients: Formulation 1: 20 mM sodium phosphate, 200 mM glycine, 0.01% polysorbate 20 (PS20), pH 6.5 Formulation 2: 50 mM sodium acetate / Na acetate salt, 200 mM glycine 0.01% PS20, pH 5.5 Formulation A: 50 mM acetate / Na acetate, 7% (w / v) sucrose, 0.01% PS20, pH 5.4 Formulation B: 50 mM acetic acid / Na acetate, 2.5% (w / v) sorbitol, 70 mM NaCl, 0.01% PS20, pH 5.4 Formulation C: 50 mM acetate / Na acetate, 100 mM arginine, 100 mM glutamate, 0.01% PS20, pH 5.4

[0101] Formulation C was selected as the planned formulation of 200 mg / mL KPL-404 for drug substance (DS) and drug product (DP) with 0.02% PS20 instead of 0.01% PS20.

[0102] Screening Tests To evaluate the stability of KPL-404 in the formulation screen, KPL-404 was exchanged into 12 different buffers, including formulation A (pH 5.6), formulation 1, and formulation 2. Freeze / thaw (F / T) stress was evaluated by cycling the samples from -70C to ambient temperature three times. Agitation stress was also evaluated by shaking at 200 rpm for 48 hours in an upright position. Short-term stability studies were performed by storing samples at 2-8°C, 25°C, or 40°C for up to 3 weeks. The SEC results indicate that excipients play a key role in ensuring the stability of KPL-404 after F / T and agitation stress, and glycine formulations 1 and 2 are susceptible to degradation, i.e., loss of % major species or increase in either % HMW species or % LMW species. KPL-404 was stable in formulation A, which contains sucrose as a stabilizer. The SEC results of the screened formulations A, 1, and 2 are graphically shown in Figure 1. There were clear differences between the formulations, with formulation 2 being the least stable, showing a loss of the major species and an increase in both HMW and LMW species. Formulation 1 showed an increase in HMW species at 40°C and was unstable during the F / T stress and agitation tests. Formulation A was the lead candidate in these screening studies and was selected for further development studies.

[0103] Development Testing Following the screening study, further studies were conducted on three candidate formulations that were identified as part of the development studies.

[0104] KPL-404 was formulated by UF / DF at 200 mg / mL in each of the three candidate formulations (Formulations AC) and stability assayed in vial presentation.

[0105] KPL-404 was highly concentrated by UF / DF until the flux was zero. The concentrations achieved were as follows: 272 mg / mL KPL-404 in formulation A: (50 mM acetate, 7% (w / v) sucrose, 0.001% PS20, pH 5.4) 274 mg / mL KPL-404 in formulation B: 50 mM acetate, 2.5% (w / v) sorbitol, 70 mM NaCl, 0.001% PS20, pH 5.4 291 mg / mL KPL-404 in formulation C: 50 mM acetate, 100 mM arginine / 100 mM glutamate, 0.001% PS20, pH 5.4

[0106] Each of these samples was characterized externally using biophysical techniques to characterize self-association as a function of protein concentration. The drug substance used to prepare these formulations was initially manufactured with 0.001% PS20. Separately, the formulation was spiked with 10% (w / v) PS20 to achieve 0.01% PS20 and was also analyzed.

[0107] In parallel, CG-MALS studies were performed on a formulation containing 0.01% PS20.

[0108] Viscosity and Instron tests were performed to assess syringeability as a function of protein concentration.

[0109] From these characterization studies, it was concluded that KPL-404 could achieve concentrations up to 292 mg / mL in Formulation C. Rapid stability studies were performed at 220 mg / mL for each of Formulations A and B, and at 235 mg / mL for Formulation C.

[0110] Formulation C at 200 mg / mL KPL-404 was selected based on improved properties compared to formulations A and B, for both drug substance and formulation.

[0111] material Material generation to evaluate high concentrations of KPL-404

[0112] Initial Testing The non-GMP drug substance was diluted into each of the three buffers and subsequently concentrated to 200 mg / mL. Polysorbate 20 present in the drug substance was expected to co-concentrate with KPL-404 in the centrifuge unit.

[0113] Material generation for stability testing UFDF Test: To generate the charge material for each ProA cycle, the drug substance was thawed in a 25° C. water bath and adjusted to a target of 10 g / L using ProA equilibration buffer. The adjusted ProA charge material was 0.2 μm filtered before loading onto the column. Each ProA eluate was neutralized to a target pH of 5.00, 0.2 μm filtered, and stored at or below −65° C. A total of four cycles were performed to generate 68 grams of UFDF charge material for high concentration testing.

[0114] Material generation for UFDF verification: To generate the charge material for each ProA cycle, the drug substance was thawed overnight at 2-8°C and adjusted to a target of 10 g / L using ProA equilibration buffer. The adjusted ProA charge material was 0.2 μm filtered prior to loading onto the column. Each ProA eluate was 0.2 μm filtered and stored at or below -65°C. A total of three cycles were performed to generate 54 grams of UFDF charge material for high concentration testing.

[0115] For each run, the neutralized ProA eluate was thawed at 2-8 °C, 0.2 μm filtered, and used as the UFDF load. The starting material was concentrated to a target of 50 g / L, diafiltered into the relevant buffer, and then concentrated until a prohibitive drop in flux occurred or the retentate showed visual degradation. During processing, when the 200 g / L target was reached, a 40 mL in-process pool was sampled for formulation processing. The remaining pool was then further concentrated while adjusting the feed flow rate and transmembrane pressure to avoid exceeding device capacity at high concentrations and pressures.

[0116] A system rinse using 1-1.5x hold-up volume was performed to allow for mass balance calculations of the process steps. A 10% w / v PS20 stock solution was prepared in each respective formulation buffer and spiked into both target 200g / L samples to a target of 0.01% PS20. A final 0.2μm filtration was performed using a Sartorius 3.5cm 2The procedure was carried out using a PES filter. Each pool was filtered separately and the pressure was monitored. The remaining drug substance pool was handled according to the formulation stability study to ensure compliance with study objectives.

[0117] Results and Discussion Effect of formulations A to C on KPL-404 at levels above 200 mg / mL Small scale UF / DF was performed to generate the following samples, which were concentrated until tangential flow ceased and were deemed to be at the threshold of solubility: Formulation A (sucrose-based, Ph1 formulation): 50 mM acetate, 7% (w / v) sucrose, 0.01% PS20, pH 5.4 - achieving 212.6 mg / mL KPL-404 Formulation B (sorbitol-based): 50 mM acetate, 2.5% (w / v) sorbitol, 70 mM NaCl, 0.01% PS20, pH 5.4 - achieving 195.5 mg / mL KPL-404 Formulation C (arginine / glutamate based): 50 mM acetate, 100 mM arginine / 100 mM glutamate, 0.01% PS20, pH 5.4 - achieving 214.1 mg / mL KPL-404

[0118] The study used KPL-404 drug substance (i.e., an antibody having a heavy chain variable region having the amino acid sequence set forth in SEQ ID NO:9 and a light chain variable region having the amino acid sequence set forth in SEQ ID NO:10) that was processed through a Protein A column to remove all remaining buffer components, including PS20, and then ultrafiltered / diafiltered (UFDF) into each of Formulation A, Formulation B, or Formulation C at 200 mg / mL. The formulations were then further concentrated to approximately 250 mg / mL. All were spiked with PS20 to a target of 0.01% and then filtered.

[0119] Biophysical characterization Malvern Panalytical Testing The formulations were tested by various biophysical techniques including dynamic light scattering, static light scattering (B22 and CIMax), electrophoretic light scattering (Zeff), thermodynamic stability (melting transition) by differential scanning calorimetry, injectability by microcapillary viscosity and rheology, and nanoparticle tracking analysis.

[0120] An initial set of formulations was characterized at 200 mg / mL as shown in Table 2. Formulation C was discovered to be devoid of PS20 after testing was completed. [Table 2] [Table 3]

[0121] Of the formulations containing trace levels of PS20, formulation C had the lowest colloidal stability based on kD values. Similarly, formulation C had the highest concentration of subvisible particles (C) compared to formulations A and B. sμp ) (Figure 2A).

[0122] Thermal analysis showed similar thermal stability for the onset of aggregation and for each of the three observed thermal transitions, with Formulation A being more stable due to sucrose compared to Formulations B and C, which were essentially equivalent.

[0123] The biophysical characterization study was repeated after the generation of higher concentration (>200 mg / mL) KPL-404 formulations, including 0.02% PS20 in formulation C, Table 3. Formulation C had the highest colloidal stability based on kD values ​​when 0.02% PS20 was in the formulation. Similarly, formulation C had the lowest colloidal stability based on kD values ​​when compared to formulation B. sμp ) were at lower concentrations (Fig. 2B ).

[0124] Rheological evaluation of viscosity as a function of shear rate showed different behavior above 100 mg / mL and 200 mg / mL (Figure 3).

[0125] At 100 mg / mL, all formulations exhibited Newtonian behavior with similar viscosities as a function of shear rate. However, above 200 mg / mL, there is a common non-Newtonian "shear thinning" effect for formulations A and B, but a statically invariant viscosity versus shear rate is observed for formulation C. This suggests that the arginine / glutamate excipients in formulation C inhibit self-association at higher concentrations, whereas there was low energy self-association at low shear rates for formulations A and B, which collapsed as shear rate increased.

[0126] This behavior is consistent with a change in the mechanism of colloidal stability above 250 mg / mL, where protein self-association becomes dominant rather than the hard-sphere mixture model that characterizes dilute protein solutions of KPL-404 in formulations A and B. Conversely, for KPL-404 in formulation C, the absence of shear thinning at high concentrations leads to an exponential increase in viscosity as a function of protein concentration up to 292 mg / mL. The combination of arginine and glutamate stabilizes KPL-404 against self-association via a different mechanism than sucrose in formulation A or sorbitol / NaCl in formulation B.

[0127] In summary, biophysical characterization of KPL-404 formulations indicates differences in the mechanisms of stabilization, with sucrose formulation A being less optimal than either formulations B or C. The data suggest that formulation C may be the most effective at limiting self-association at high concentrations through a combination of arginine / glutamate and PS20.

[0128] Biophysical Testing Concentration gradient multi-angle light scattering (CG-MALS) was used to assess the oligomeric behavior of KPL-404 over a wide range of concentrations. Individual samples were prepared at different concentrations and analyzed by static light scattering (SLS).

[0129] Wyatt technologies implemented CG-MALS by incorporating an autosampler that generates a concentration gradient from two solutions (from a high-concentration protein formulation and a matched placebo) in conjunction with in-line sample analysis.

[0130] KPL-404 formulations and compatible formulation buffers were filtered using a 0.22 μm syringe filter. Fifteen concentrations per formulation were analyzed and concentrations were confirmed by UV absorbance.

[0131] Fitting of the data to a reversible / irreversible association model was performed using Wyatt Calypso software (144 kDa monomer).

[0132] CG-MALS results for formulation A

[0133] It was not possible to model reversible association at the highest concentration included.

[0134] The inability to approximate reversible oligomers may be due to repulsive interactions at higher concentrations that dominate and mask the underlying attractive interactions.

[0135] Due to strong repulsive interactions that masked the underlying attractive interactions, the highest concentration at which the reversible association model could be approximated was approximately 127 mg / mL for the reversible dimer.

[0136] The data up to about 127 mg / mL can be modeled with a reversible dimer. The concentration of the reversible dimer did not exceed that of the monomer within the concentration range that could be approximated. The reversible dimer was present even at the lowest concentration (about 0.5 mg / mL). See Figure 4.

[0137] CG-MALS results for formulation B

[0138] Similarly, it was not possible to model reversible association at the highest concentration points included: due to strong repulsive interactions that masked the underlying attractive interactions, the highest concentration at which a reversible association model could be fitted was approximately 163 mg / mL for the reversible dimer and hexamer (Figure 5).

[0139] The proportion of reversible oligomers exceeds that of monomers at approximately 35 mg / mL. Reversible dimers were present even at the lowest concentrations (approximately 0.5 mg / mL).

[0140] CG-MALS results for formulation C

[0141] Similarly, it was not possible to model reversible association at the highest concentration points included: due to strong repulsive interactions that masked the underlying attractive interactions, the highest concentration at which a reversible association model could be fitted was approximately 126 mg / mL for the reversible dimer and hexamer (Figure 6).

[0142] The proportion of reversible oligomers exceeds that of monomers at about 60 mg / mL. Reversible hexamers begin to form at about 25 mg / mL. Even at the lowest total protein concentration, about 2% dimers were present.

[0143] In summary, CG-MALS shows that KPL-404 readily forms reversible dimers and higher oligomers that are not detected by SEC (diluted to approximately 1 mg / mL prior to injection).

[0144] The results were generally consistent with hard-ball repulsive behavior, with underlying attractive behavior in all formulations as indicated by the need to include reversible associations in the global analysis.

[0145] Formulation A was more effective at inhibiting the formation of reversible oligomers below 125 mg / mL, but the data were not comparable at higher concentrations.

[0146] Formulation B was the least stable with respect to oligomerization, with reversible oligomers becoming the predominant species at approximately 35 mg / mL.

[0147] Formulation C showed that oligomers were the predominant species above 60 mg / mL and had a lower viscosity compared to Formulation A, which is consistent with Formulation C having less energetic self-association at higher concentrations.

[0148] These data suggest that KPL-404 is unlikely to aggregate not only at concentrations above 200 mg / mL, but also at lower concentrations, e.g., 150-200 mg / mL. Both rheology and light scattering data suggest that KPL-404 self-association tends to increase with increasing concentration. Shear thinning data reveals a striking and unexpected difference in the self-association kinetics of formulation C from the other formulations, suggesting that arginine and glutamate affect these interactions through different mechanisms. Light scattering data indicate that in formulations A, B, and C below approximately 100 mg / mL, KPL-404 is primarily monomeric. However, at higher concentrations, these data suggest that formulation C has a superior ratio of reversible to irreversible dimers compared to formulations A and B.

[0149] Characterization of needle penetration. Viscosity was evaluated as a function of varying concentration for each of the three formulations, Table 4 and FIG.

[0150] Viscosity varied exponentially with protein concentration. The inset in Figure 7 highlights the KPL-404 concentration at which a given formulation exceeds 25 cP, the theoretical upper limit for syringeability based on extrusion force. Formulation A with sucrose exceeds 25 cP at approximately 200 mg / mL, formulations B and C exceed 25 cP at over 220 mg / mL, and formulation C is slightly less viscous than B. [Table 4]

[0151] Each of formulations A, B, and C was hand-filled into a 2.25 mL BD Neopak syringe. The same sample was tested five times with a new syringe for each test. Instron testing was performed to record the sliding force as a function of protein concentration for each formulation (Table 5). To set aside samples, the test utilized the highest concentration sample of each formulation and the collected samples were diluted with buffer to generate lower concentrations. The results show a decrease in ejection force as a function of decreasing concentration.

[0152] Using a manually assembled autoinjector (AI) with each of the candidate formulations in a 2.25 mL BD Neopak syringe, the injection times for the three lowest concentrations were recorded, Table 5. Times were established by calculating the time from the video recording, frame rate and number of frames from the first to last drop.

[0153] The standard Ypsomate AI was tested on all three candidates: Injection times (into air) for Formulation A were greater than 20 seconds at 220 mg / mL and greater than 15 seconds at 200 mg / mL. Injection time for Formulation B was over 20 seconds for 237 mg / mL and 15 seconds for 220 mg / mL. Injection times for Formulation C were greater than 20 seconds for 252 mg / mL and 15 seconds for 234 mg / mL. Ypsomate Pro AI was tested only on Formulation C: Injection times for Formulation C were greater than 20 seconds for 271 mg / mL and 15 seconds for 262 mg / mL. See Table 5 and Figures 8-10B. [Table 5]

[0154] In summary, sliding force and injection time generally correlate with viscosity, but formulation C consistently achieved higher concentrations compared to formulation B, and sucrose formulation A performed less well than B and C.

[0155] Stability of KPL-404 Formulations at 200 mg / mL and Greater than 200 mg / mL All three formulations were examined for stability at 200 mg / mL, the upper concentration limit for Formulation A based on injectability data.

[0156] Injectability data suggests that Formulation B can support 220 mg / mL. For direct comparison, both Formulations A and B were subjected to short-term stability (3 months) at 220 mg / mL. Injectability data suggests that Formulation C can support 235 mg / mL, which was also subjected to short-term stability (3 months). The data are summarized in Tables 6-14. [Table 6] [Table 7] [Table 8] [Table 9] [Table 10] [Table 11] [Table 12] [Table 13] [Table 14]

[0157] Subsequent aggregation data was generated for Formulation A (100 mg / mL KPL-404 in 50 mM sodium acetate, 7% (w / v) sucrose, 0.01% (w / v) polysorbate 20, pH 5.4) KPL-404 at 2-8 °C for up to 24 months. For the first lot, the SEC major peak was 98.3% at T = 0, 97.6% at T = 12 months and T = 24 months, indicating a ΔSEC (major peak) of -0.71%, which did not change from 12 months to 24 months. The SEC high molecular weight (HMW) peak, representing antibody aggregates, was 1.3% at T = 0, 1.6% at T = 12 months, and 1.8% at T = 24 months, indicating little increase in aggregation over time. For the second lot, the SEC major peak was 98.6% at T=0 and 97.8% at T=12 months, giving a ΔSEC (major peak) of -0.81%. Data was not generated until 24 months. The SEC high molecular weight (HMW) peak for the second lot was 0.9% at T=0, 1.4% at T=12 months, and 1.5% at T=18 months, again indicating little increase in aggregation over time.

[0158] Aggregation data was also generated for formulation C (200 mg / mL KPL-404 in 50 mM sodium acetate, 100 mM L-arginine, 100 mM L-glutamate, 0.02% polysorbate 20, pH 5.4) KPL-404 at 2-8 °C for up to 12 months. In the first lot, the protein showed SEC major peaks of 99.2% and 98.9% at T=0 and T=12 months, respectively, with a ΔSEC (major peak) of -0.3%. The HMW peak was 0.60% at T=0 and 0.70% at T=12 months, indicating a slight increase in aggregation over time. In the second lot, the protein showed SEC major peaks of 98.7% and 98.1% at T=0 and T=9 months, respectively, with a ΔSEC (major peak) of -0.61%. The HMW peak was 0.40% at T=0 and 0.70% at T=9 months, indicating a slight increase in aggregation over time.

[0159] Aggregation by SEC is the major degradation pathway. Figure 1 shows a set of graphs illustrating the SEC results plotted for % major, % HMW, and % LMW species of KPL-404 for formulations A, 1, and 2. The chromatograms of the initial samples are shown in Figure 11 and show that initially all three formulations are essentially the same as the reference standard.

[0160] After two weeks of heat stress at 40° C., FIG. 12, aggregation was observed for all three formulations, with the formation of species larger than dimers for formulations A and B. Formulation C, in comparison, minimized the formation of higher order species, consistent with the arginine / glutamate excipients allowing for higher protein concentrations of KPL-404 compared to sucrose and sorbitol.

[0161] Statistical analysis was performed on SEC results from formulations held at recommended storage conditions, 5°C (Figure 13), accelerated storage conditions, 25°C (Figure 14), stressed storage conditions, 40°C (Figure 15), and frozen storage conditions, -70°C (F6).

[0162] Statistical analysis assessed the null hypothesis (zero slope) with one-sided 95% confidence intervals. If the slope was not statistically zero, statistical significance was set at a p-value <0.05, Table 15.

[0163] KPL-404 is stable at 5°C in the current formulation at 100 mg / mL, showing a slow rate of change over 12 months with a slope predicting approximately 1% change after 60 months. Of the three formulations at 200 mg / mL, formulation C is more stable than both formulations A and B, with the slope of formulation C predicting approximately 1% change after approximately 36 months, while A and B predict twice the rate of change of formulation C. At 220 mg / mL for A and B and 235 mg / mL for C, there is a sharp increase in the primary loss through aggregation for all three formulations.

[0164] The trends observed at 5°C were reproduced under accelerated stress conditions at 25°C, with major % changes occurring at similar rates for Formulation A at 100 mg / mL and Formulation C at 200 mg / mL. The stability of KPL-404 in Formulation C at 235 mg / mL was essentially equivalent at 5°C, while being consistently more stable than Formulations A and B at 220 mg / mL.

[0165] The trends observed at 40°C mirrored those at 25°C and 5°C, with the major % changes occurring at similar rates for Formulation A at 100 mg / mL and Formulation C at 200 mg / mL. Note that only Formulation A at 100 mg / mL had sufficient data for meaningful statistical analysis. Analysis of the higher concentration data set is included in Table 15 for qualitative comparison, which shows that Formulation C was superior to Formulations A and B in reducing aggregation when held at stressed storage conditions of 40°C. The results also indicate that KPL-404 is a monoclonal antibody that is generally stable against aggregation, even at the high concentrations tested here.

[0166] KPL-404 was stable at -70°C with changes in % major occurring at similar rates for Formulation A at 100 mg / mL and Formulations A, B, and C at 200 mg / mL. Note that only Formulation A at 100 mg / mL had sufficient data for meaningful statistical analysis. Analysis of the higher concentration data sets is included in Table 15 for qualitative comparison. The data predicts no statistically significant changes in % major purity during frozen storage over the 12 month data. [Table 15]

[0167] In summary, KPL-404 is stable in all three formulations, but formulation C is superior to both formulations A and B in supporting concentrations above 200 mg / mL.

[0168] Studies were conducted with three candidate formulations. KPL-404 was formulated by UF / DF at 200 mg / mL in each of the three candidate formulations and stability was established in vial presentation.

[0169] KPL-404 was highly concentrated by UF / DF until the flux reached zero. The concentrations achieved were 272 and 274 mg / mL for formulations A and B, respectively, and 291 mg / mL for formulation C.

[0170] Each of these samples was characterized externally using biophysical techniques available at Malvern Panalytical and FDB.

[0171] These studies indicate that KPL-404 is well represented by a reversible monomer-dimer equilibrium below about 130 mg / mL.

[0172] At higher concentrations, approximately 130-200 mg / mL, higher order reversible oligomers were formed to varying degrees in all three formulations.

[0173] Viscosity and Instron tests were performed to evaluate syringeability as a function of protein concentration. Results showed that formulation C was more effective in reducing viscosity and could support higher concentrations compared to formulations A and B.

[0174] Formulation C shows excellent properties with either the standard Yposmate autoinjector at up to 235mg / mL or the Yposmate Pro at up to 270mg / mL.

[0175] All three formulations were subjected to stability at 200 mg / mL, the upper concentration limit for Formulation A based on injectability data. Short-term (3 month) stability was also evaluated at 220 mg / mL for Formulations A and B, and at 235 mg / mL for Formulation C.

[0176] Formulation C at 200 mg / mL was almost as stable as Formulation A at 100 mg / mL.

[0177] Based on all the results collected, Formulation C was identified as the superior formulation for KPL-404 compared to Formulations A and B. For the final UF / DF of the drug substance, an upper concentration limit of 250 mg / mL can be supported. EXAMPLES

[0178] A 26-week repeat-dose toxicity and toxicokinetic study of KPL-404 administered intravenously or subcutaneously once weekly to cynomolgus monkeys with an 8-week recovery period

[0179] The objective of this study was to evaluate the toxicity of KPL-404 prepared in Formulation C (200 mg / mL KPL-404, 50 mM acetate, 100 mM L-arginine, 100 mM L-glutamate, 0.02% polysorbate 20, pH 5.4) when administered by intravenous or subcutaneous injection to cynomolgus monkeys once a week for a period of 26 weeks, and to evaluate the potential reversibility or delayed onset of any effects over an 8-week recovery period. In addition, the toxicokinetic properties of the test article were determined. KPL-404 was administered by subcutaneous (SC) or intravenous bolus (IV) injection to 5 male and 5 female cynomolgus monkeys per group at doses of 30 or 97 mg / kg once a week for 26 weeks according to the study design in Table 1 of the paper. The control substance, Formulation C, without KPL-404, was administered by SC injection at the same dosing frequency. [Table 16]

[0180] Toxicology results No unexpected adverse events were observed in cynomolgus monkeys treated with placebo or either the low dose of 30 mg / kg or the high dose of 97 mg / kg of KPL-404 in Formulation C. All observed effects were attributable to the pharmacology of KPL-404.

[0181] conclusion KPL-404 was stable in all three formulations, however, formulation C was superior to both formulations A and B in providing a stable formulation of KPL-404, and formulation C was further demonstrated to be safe in non-human primate toxicity studies.

[0182] Other embodiments Although certain aspects of the present invention have been described and illustrated, such aspects are merely illustrative of the present invention and should not be construed as limiting the present invention as interpreted according to the appended claims. All publications and patent applications cited herein are incorporated by reference in their entirety for all purposes as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference for all purposes. Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it will be readily apparent to those skilled in the art in light of the teachings of the present invention that certain changes and modifications can be made without departing from the spirit or scope of the appended claims.

Claims

1. A pharmaceutical composition comprising a humanized anti-CD40 antibody or antigen-binding fragment thereof, wherein the antibody comprises a heavy chain variable region comprising the amino acid sequence set forth in SEQ ID NO:9 and a light chain variable region comprising the amino acid sequence set forth in SEQ ID NO:10, wherein the pharmaceutical composition comprises polysorbate 20, acetate, and a polar excipient.

2. 10. The pharmaceutical composition of claim 1, wherein the pharmaceutical composition comprises about 5 mM to about 100 mM acetate, optionally wherein the pharmaceutical composition comprises about 50 mM acetate.

3. 10. The pharmaceutical composition of claim 1, wherein the pharmaceutical composition comprises about 0.001% to about 0.1% polysorbate 20, optionally wherein the pharmaceutical composition comprises about 0.02% polysorbate 20 or about 0.01% polysorbate 20.

4. 10. The pharmaceutical composition of claim 1, wherein the pharmaceutical composition has a pH of about 5.0 to about 6.0, and optionally, the pharmaceutical composition has a pH of about 5.

4.

5. 10. The pharmaceutical composition of claim 1, wherein the polar excipient comprises arginine and glutamate.

6. 6. The pharmaceutical composition of claim 5, wherein the pharmaceutical composition comprises about 1 mM to about 200 mM arginine and / or about 1 mM to about 200 mM glutamate, optionally wherein the pharmaceutical composition comprises about 100 mM arginine and / or about 100 mM glutamate.

7. 6. The pharmaceutical composition of claim 5, wherein the pharmaceutical composition comprises about 0.001% to about 0.1% polysorbate 20, optionally wherein the pharmaceutical composition comprises about 0.01% polysorbate 20 or about 0.02% polysorbate 20.

8. 6. The pharmaceutical composition of claim 5, wherein the pharmaceutical composition comprises about 50 mM acetate, about 100 mM arginine, about 100 mM glutamate, and about 0.02% polysorbate 20.

9. 10. The pharmaceutical composition of claim 1, wherein the polar excipient comprises sucrose, and optionally the pharmaceutical composition comprises from about 1% (w / v) to about 10% (w / v) sucrose.

10. 10. The pharmaceutical composition of claim 9, wherein the pharmaceutical composition comprises about 7% (w / v) sucrose.

11. 10. The pharmaceutical composition of claim 9, wherein the pharmaceutical composition comprises about 50 mM acetate, about 7% (w / v) sucrose, and about 0.02% or about 0.01% polysorbate 20.

12. 10. The pharmaceutical composition of claim 1, wherein the polar excipient comprises sorbitol, and optionally the pharmaceutical composition comprises from about 0.1% (w / v) sorbitol to about 5% (w / v) sorbitol.

13. 13. The pharmaceutical composition of claim 12, further comprising sodium chloride, optionally wherein the pharmaceutical composition comprises from about 1 mM to about 100 mM sodium chloride.

14. 14. The pharmaceutical composition of claim 13, wherein the pharmaceutical composition comprises about 70 mM sodium chloride.

15. 13. The pharmaceutical composition of claim 12, wherein the pharmaceutical composition comprises about 2.5% (w / v) sorbitol.

16. 14. The pharmaceutical composition of claim 13, wherein the pharmaceutical composition comprises about 50 mM acetate, about 2.5% (w / v) sorbitol, about 70 mM NaCl, and about 0.02% or about 0.01% polysorbate 20.

17. 10. The pharmaceutical composition of claim 1, wherein the antibody or antigen-binding fragment thereof is at a concentration of about 50 mg / mL to about 300 mg / mL, optionally wherein the antibody or antigen-binding fragment thereof is at a concentration of about 200 mg / mL.

18. (a) the humanized anti-CD40 antibody or antigen-binding fragment thereof is KPL-404; (b) at least 95% or at least 98% of the humanized anti-CD40 antibody or antigen-binding fragment thereof is present in the composition in monomeric form, and / or 95% or more or 98% or more of the humanized anti-CD40 antibody or antigen-binding fragment thereof is present in the composition in monomeric form after 12 months of storage at 2-8°C, and / or (c) the pharmaceutical composition of claim 1, wherein less than 5% or less than 2% of the humanized anti-CD40 antibody or antigen-binding fragment thereof is present in the composition as high molecular weight (HMW) species and / or less than 5% or less than 2% of the humanized anti-CD40 antibody or antigen-binding fragment thereof is present in the composition as high molecular weight (HMW) species after storage at 2-8°C for 12 months.

19. A kit comprising the pharmaceutical composition of any one of claims 1 to 18 and instructions for its use.

20. A pharmaceutical composition according to any one of claims 1 to 18 for use in suppressing the immune system of a human subject.

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