Anti-FGFR2 antibody formulations

Abstract

To provide formulations of anti-fibroblast growth factor receptor 2 (FGFR2) antibody that can be stored in a liquid ready-to-use form and can be intravenously administered by intravenous (IV) infusion.SOLUTION: Provided herein is a pharmaceutical formulation comprising: i) 10 to 30 mg / mL of an anti-fibroblast growth factor receptor 2 (FGFR2) antibody; ii) 5 to 40 mM of a buffer solution selected from one or more of histidine, citric acid or phosphate; iii) 130 to 170 mM arginine or 250-290 mM sucrose; iv) 0.002% to 0.1% polysorbate 20 or polysorbate 80, where the formulation has a pH of 5.0 to 6.5.SELECTED DRAWING: None

Description

[Technical field]

[0001] This application claims the benefit of priority to U.S. Provisional Patent Application No. 62 / 741,772, filed October 5, 2018, which is incorporated by reference in its entirety.

[0002] The present disclosure relates to formulations for anti-FGFR2 (fibroblast growth factor receptor 2) antibodies, such as anti-FGFR2-IIIb antibodies, which in some embodiments can be stored for extended periods of time as a liquid, e.g., in a ready-to-use form, and in some embodiments can be administered intravenously, e.g., by intravenous (IV) infusion. [Background technology]

[0003] The present disclosure relates to specific formulations of antibodies that bind to FGFR2, such as the splice form FGFR2-IIIb (fibroblast growth factor receptor 2 IIIb). FGFR2 is one of four FGFR proteins (FGFR1, 2, 3, and 4). FGFRs are characterized by multiple alternative splicing of their mRNAs, resulting in various isoforms (Ornitz et al., J. Biol. Chem. 271:15292, 1996, see also Swiss-Prot P21802 and isoforms P21802-1 to -20 for the sequence of FGFR2 and its isoforms). With respect to FGFR2, alternative splicing results in, for example, the isoforms FGFR2-IIIb and FGFR2-IIIc (or simply FGFR2b and FGFR2c). The FGFR2-IIIb form of FGFR2 (also called K-sam-II) is a high affinity receptor for both FGF1 and KGF family members (FGF7, FGF10, and FGF22), while FGFR2-IIIc (also called K-sam-I) binds well to both FGF1 and FGF2, but not to KGF family members (Miki et al., Proc. Natl. Acad. Sci. USA 89:246, 1992). FGFR2-IIIb is the only receptor for KGF family members (Ornitz et al., 1996, ibid.), and is therefore also called KGFR.

[0004] The inhibitor of FGFR2 can include antibodies. For example, US Patent No. 8,101,723B2 describes a monoclonal antibody that binds to human FGFR2-IIIb, but binds less well or does not bind to FGFR2-IIIc, and vice versa. US Patent Publication No. 2015-0050273A1 describes a certain afucosylated antibody that binds to FGFR2-IIIb. The present disclosure describes formulations that can be used for the antibodies described in, for example, US Patent No. 8,101,723B2 and US Patent Publication No. 2015-0050273A1.

[0005] In some embodiments, formulation conditions are specified herein that allow the anti-FGFR2 antibodies described herein to be prepared in the form of a liquid formulation that remains stable after several months of storage and can be administered intravenously, for example by IV infusion. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] U.S. Pat. No. 8,101,723 [Patent Document 2] US Patent Application Publication No. 2015 / 0050273 [Non-patent literature]

[0007] [Non-Patent Document 1] Ornitz et al.,J.Biol.Chem.271:15292,1996 [Non-Patent Document 2] Miki et al.,Proc.Natl.Acad.Sci.USA 89:246,1992 Summary of the Invention

[0008] The disclosure encompasses, for example, pharmaceutical formulations of anti-FGFR2 antibodies comprising: i) 10-30 mg / mL antibody; ii) 5-40 mM buffer selected from one or more of histidine, citrate, or phosphate; iii) 130-170 mM arginine or 250-290 mM sucrose; and iv) 0.002%-0.1% polysorbate 20 or polysorbate 80, wherein the formulation has a pH of 5.0-6.5; and the anti-FGFR2 antibody is an antibody comprising: a) a heavy chain comprising a sequence of SEQ ID NO:2 and a light chain comprising a sequence of SEQ ID NO:3; b) a heavy chain comprising a heavy chain (HC) hypervariable region 1 (HVR1) comprising a sequence of SEQ ID NO:6, a HC HVR2 comprising a sequence of SEQ ID NO:7, and a HC HVR3 comprising a sequence of SEQ ID NO:8, and a light chain (LC) HVR1 comprising a sequence of SEQ ID NO:9, a LC HVR2 comprising a sequence of SEQ ID NO:10, and a LC HVR3 comprising a sequence of SEQ ID NO:11. and c) an antibody comprising a heavy chain comprising a variable region sequence comprising the sequence of SEQ ID NO: 4, and a light chain comprising a light chain variable region sequence comprising the sequence of SEQ ID NO: 5. In some embodiments, the formulation has one or more of the following properties: (a) is a ready-to-use liquid formulation, (b) is not lyophilized prior to administration to a patient, (c) is contained in a single-use vial, (d) protein aggregation in the formulation does not increase by more than 2.0% after six months of storage at 5° C., (e) protein aggregation in the formulation does not increase by more than 2.0% or 2.5% after three months of storage at 25° C., (f) protein aggregation in the formulation does not increase by more than 7.0% after three months of storage at 40° C. (g) the charged protein variants in the formulation do not change by more than 5% after storage for six months at 5° C.; (h) protein aggregation in the formulation does not increase by more than 2.0% after five freeze-thaw cycles at −70° C.; (i) protein aggregation in the formulation does not increase by more than 2.0% after mechanical stress at 500 rpm for 72 hours; (j) is diluted in saline solution prior to intravenous administration; (k) is administered intravenously (e.g., by intravenous infusion); and (l) is isotonic with human plasma.

[0009] In some cases, the formulation contains 10-15 mg / mL, 15-20 mg / mL, 20-25 mg / mL, 18-22 mg / mL, 10 mg / mL, 11 mg / mL, 12 mg / mL, 13 mg / mL, 14 mg / mL, 15 mg / mL, 16 mg / mL, 17 mg / mL, 18 mg / mL, 19 mg / mL, 20 mg / mL, 21 mg / mL, 22 mg / mL, 23 mg / mL, 24 mg / mL, or 25 mg / mL of anti-FGFR2 antibody. In some cases, the formulation comprises a 10-40 mM, 10-30 mM, 15-25 mM, 10-20 mM, 20-30 mM, 18-22 mM, 10 mM, 15 mM, 16 mM, 17 mM, 18 mM, 19 mM, 20 mM, 21 mM, 22 mM, 23 mM, 24 mM, 25 mM, 26 mM, 27 mM, 28 mM, 29 mM, or 30 mM buffer. In some embodiments, the buffer is a histidine buffer. In some embodiments, the formulation comprises 130-150 mM, 150-170 mM, 140-160 mM, 130 mM, 135 mM, 140 mM, 145 mM, 150 mM, 155 mM, 160 mM, 165 mM, or 170 mM arginine. In some cases, the formulation comprising arginine does not comprise sucrose. In some embodiments, the pH of the formulation comprising arginine is 5.0-7.0, 5.0-6.0, 5.5-6.0, 5.5-6.5, 5.5-5.9, 5.6-5.8, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, or 6.5. In some cases, the pH of the formulation is 5.5-5.9, 5.6-5.8, 5.6, 5.7, 5.8, 5.9, or 6.0. In some embodiments, the formulation comprises 260-280 mM, 250 mM, 260 mM, 270 mM, 280 mM, 290 mM sucrose. In some such cases, the formulation comprising sucrose does not comprise arginine. In some embodiments, the pH of the formulation comprising sucrose is 5.0-7.0, 5.0-6.0, 5.5-6.0, 5.5-5.9, 5.6-5.8, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, or 6.5.In some embodiments, the pH of the formulation is 5.8-6.2, 5.9-6.1, 5.9, 6.0, or 6.1.

[0010] Any of the above formulations may also contain 0.01-0.1%, 0.005-0.05%, 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 or 80. In some cases, the formulation contains 0.01-0.1%, 0.005-0.05%, 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.

[0011] In some embodiments, the formulation consists essentially of an anti-FGFR2 antibody; a citrate, phosphate, or histidine buffer; arginine or sucrose; and polysorbate 20 or 80. In some such cases, the formulation consists essentially of an anti-FGFR2 antibody, histidine, arginine or sucrose, and polysorbate 20.

[0012] The present disclosure also includes, for example, a pharmaceutical formulation of an anti-FGFR2 antibody, the formulation being a liquid formulation that is not lyophilized prior to use, comprising i) 10-25 mg / mL of an anti-FGFR2 antibody, ii) 10-30 mM histidine buffer, iii) 140-160 mM arginine, and iv) 0.005%-0.05% polysorbate 20, the formulation having a pH of 5.6-5.8. The present disclosure also includes, for example, a pharmaceutical formulation of an anti-FGFR2 antibody, the formulation being a liquid formulation that is not lyophilized prior to use, comprising i) 10-25 mg / mL of an anti-FGFR2 antibody, ii) 20 mM histidine buffer, iii) 150 mM arginine, and iv) 0.01% polysorbate 20, the formulation having a pH of 5.7. Embodiments herein further include a pharmaceutical formulation of an anti-fibroblast growth factor receptor 2 (FGFR2) antibody comprising: i) 20 mg / mL anti-FGFR2 antibody; ii) 20 mM L-histidine; iii) 150 mM L-arginine; and iv) 0.01% polysorbate 20, the pharmaceutical formulation having a pH of 5.7 and being a liquid formulation that is not lyophilized prior to use. In any of these cases, in some embodiments, the formulation consists essentially of the anti-FGFR2 antibody, histidine, arginine, and polysorbate 20. In some such embodiments, the anti-FGFR2 antibody may be selected from: a) an antibody comprising a heavy chain comprising the sequence of SEQ ID NO:2 and a light chain comprising the sequence of SEQ ID NO:3; b) an antibody comprising a heavy chain comprising a heavy chain (HC) hypervariable region 1 (HVR1) comprising the sequence of SEQ ID NO:6, a HC HVR2 comprising the sequence of SEQ ID NO:7, and a HC HVR3 comprising the sequence of SEQ ID NO:8, and a light chain comprising a light chain (LC) HVR1 comprising the sequence of SEQ ID NO:9, a LC HVR2 comprising the sequence of SEQ ID NO:10, and a LC HVR3 comprising the sequence of SEQ ID NO:11; and c) an antibody comprising a heavy chain comprising a variable region sequence comprising the sequence of SEQ ID NO:4, and a light chain comprising a light chain variable region sequence comprising the sequence of SEQ ID NO:5.The pharmaceutical formulation may have one or more of the following properties: (a) it is contained in a single-use vial; (b) protein aggregation in the formulation does not increase by more than 2.0% after six months of storage at 5° C.; (c) protein aggregation in the formulation does not increase by more than 2.0% or 2.5% after three months of storage at 25° C.; (d) protein aggregation in the formulation does not increase by more than 7.0% after three months of storage at 40° C.; (e) the charged protein variants in the formulation do not change by more than 5% after six months of storage at 5° C.; (f) protein aggregation in the formulation does not increase by more than 2.0% after five freeze-thaw cycles at −70° C.; (g) protein aggregation in the formulation does not increase by more than 2.0% after 72 hours of mechanical stress at 500 rpm; (h) it is diluted in saline solution prior to intravenous administration; (i) it is administered intravenously (e.g., by intravenous infusion); and (j) it is isotonic with human plasma. The above pharmaceutical formulations containing 10 to 25 mg / mL of antibody may more specifically contain 10 to 15 mg / mL, 15 to 20 mg / mL, 20 to 25 mg / mL, 18 to 22 mg / mL, 10 mg / mL, 11 mg / mL, 12 mg / mL, 13 mg / mL, 14 mg / mL, 15 mg / mL, 16 mg / mL, 17 mg / mL, 18 mg / mL, 19 mg / mL, 20 mg / mL, 21 mg / mL, 22 mg / mL, 23 mg / mL, 24 mg / mL, or 25 mg / mL of anti-FGFR2 antibody.

[0013] The disclosure also includes, for example, a pharmaceutical formulation of an anti-FGFR2 antibody, the formulation being a liquid formulation that is not lyophilized prior to use, comprising i) 10-25 mg / mL of an anti-FGFR2 antibody, ii) 10-30 mM histidine buffer, iii) 260-280 mM sucrose, and iv) 0.005%-0.05% polysorbate 20, the formulation having a pH of 5.8-6.2. The disclosure also includes, for example, a pharmaceutical formulation of an anti-FGFR2 antibody, the formulation being a liquid formulation that is not lyophilized prior to use, comprising i) 10-25 mg / mL of an anti-FGFR2 antibody, ii) 20 mM histidine buffer, iii) 270 mM sucrose, and iv) 0.01% polysorbate 20, the formulation having a pH of 6.0. In further embodiments, the pharmaceutical formulation may comprise i) 20 mg / mL anti-FGFR2 antibody, ii) 20 mM L-histidine, iii) 270 mM sucrose, and iv) 0.01% polysorbate 20, the formulation having a pH of 6.0 and being a liquid formulation that is not lyophilized prior to use. In some such embodiments, the formulation consists essentially of anti-FGFR2 antibody, histidine, sucrose, and polysorbate 20. In some such embodiments, the anti-FGFR2 antibody may be selected from: a) an antibody comprising a heavy chain comprising the sequence of SEQ ID NO:2 and a light chain comprising the sequence of SEQ ID NO:3; b) an antibody comprising a heavy chain comprising a heavy chain (HC) hypervariable region 1 (HVR1) comprising the sequence of SEQ ID NO:6, a HC HVR2 comprising the sequence of SEQ ID NO:7, and a HC HVR3 comprising the sequence of SEQ ID NO:8, and a light chain comprising a light chain (LC) HVR1 comprising the sequence of SEQ ID NO:9, a LC HVR2 comprising the sequence of SEQ ID NO:10, and a LC HVR3 comprising the sequence of SEQ ID NO:11; and c) an antibody comprising a heavy chain comprising a variable region sequence comprising the sequence of SEQ ID NO:4, and a light chain comprising a light chain variable region sequence comprising the sequence of SEQ ID NO:5.The pharmaceutical formulation may have one or more of the following properties: (a) it is contained in a single-use vial; (b) protein aggregation in the formulation does not increase by more than 2.0% after six months of storage at 5° C.; (c) protein aggregation in the formulation does not increase by more than 2.0% or 2.5% after three months of storage at 25° C.; (d) protein aggregation in the formulation does not increase by more than 7.0% after three months of storage at 40° C.; (e) the charged protein variants in the formulation do not change by more than 5% after six months of storage at 5° C.; (f) protein aggregation in the formulation does not increase by more than 2.0% after five freeze-thaw cycles at −70° C.; (g) protein aggregation in the formulation does not increase by more than 2.0% after 72 hours of mechanical stress at 500 rpm; (h) it is diluted in saline solution prior to intravenous administration; (i) it is administered intravenously (e.g., by intravenous infusion); and (j) it is isotonic with human plasma. The above pharmaceutical formulations containing 10 to 25 mg / mL of antibody may more specifically contain 10 to 15 mg / mL, 15 to 20 mg / mL, 20 to 25 mg / mL, 18 to 22 mg / mL, 10 mg / mL, 11 mg / mL, 12 mg / mL, 13 mg / mL, 14 mg / mL, 15 mg / mL, 16 mg / mL, 17 mg / mL, 18 mg / mL, 19 mg / mL, 20 mg / mL, 21 mg / mL, 22 mg / mL, 23 mg / mL, 24 mg / mL, or 25 mg / mL of anti-FGFR2 antibody.

[0014] In some embodiments of the present disclosure, the pharmaceutical formulation does not contain sugars other than sucrose, sugar alcohols, protein species other than anti-FGFR2 antibodies, surfactants other than polysorbate 20 or polysorbate 80, amino acids other than arginine and histidine, Cu 2+ , Mg 2+ , and Mn 2+ In any embodiment of the present disclosure, the formulation may be contained in a single-use vial.

[0015] The disclosure also encompasses a method of treating a solid tumor in a patient in need thereof, comprising administering to the patient an effective amount of a pharmaceutical formulation herein. In some embodiments, the formulation is administered to the patient intravenously (e.g., by intravenous infusion).

[0016] In any of the formulations herein, the anti-FGFR2 antibody may be optionally afucosylated. In any of the formulations herein, the anti-FGFR2 antibody may be a full-length antibody, or may be an Fv, single chain Fv (scFv), Fab, Fab', or (Fab') 2 In any of the formulations herein, the antibody may be chimeric, humanized, or human. In any of the formulations herein, the antibody may be bispecific, multispecific, or conjugated. [Brief description of the drawings]

[0017] [Figure 1] Average binding of afucosylated anti-FGFR2IIIb antibodies to FGFR2IIIb or Protein A in a Biacore® assay after incubation of the antibodies with different percent hydrogen peroxide concentrations (0.01%, 0.1%, and 1.0% as well as 0% (control)). Incubation of anti-FGFR2IIIb antibodies with hydrogen peroxide did not interfere with binding to FGFR2IIIb, but did interfere to some extent with binding to Protein A. [Diagram 2] 1 shows differential scanning calorimetry (DSC) thermograms of citrate / NaCl formulations of anti-FGFR2IIIb antibody at pH 4.0, 5.0, and 6.0. The data show that the development temperature of the formulations in this range increases with pH. [Diagram 3] 1 shows size-exclusion high performance liquid chromatography SE-HPLC chromatograms of anti-FGFR2IIIb antibody screening samples at pH 4, 5, 6, 7, and 8, as discussed in Example 5 below. [Figure 4]Figure 1 shows the effect of buffer pH (in citrate or phosphate NaCl buffers) on aggregate formation as determined by SE-HPLC in the pH range 4-8 and over storage periods of 0-2 months at 40° C. The figure shows a large % increase in aggregates at pH 4 compared to the other pHs. [Diagram 5] Figure 1 shows the effect of buffer pH (in citrate or phosphate NaCl buffers) on clip (fragment) formation as determined by SE-HPLC over the pH range 4-8 and over storage periods of 0-2 months at 40° C. The figure shows a large % increase in clip formation at pH 4 compared to other pHs. [Figure 6] The figure shows the effect of buffer pH (in citrate or phosphate NaCl buffers) on aggregate formation as determined by SE-HPLC over the pH range of 4-8 and over storage periods of 0-2 months at 25° C. The figure shows that all formulations generally contained less than 3.0% aggregates over this period, with the exception of the pH 8 formulation, which increased to about 2.5% to about 4% over 2 months of storage. [Figure 7] The effect of buffer pH (in citrate or phosphate NaCl buffers) on clip (fragment) formation as determined by SE-HPLC over the pH range of 4-8 and over storage periods of 0-2 months at 25° C. The figure shows an increase to nearly 1.5% of clips at pH 4 compared to the other pHs where clips remained below 0.5%. [Figure 8] Representative weak cation exchange (WCX)-HPLC chromatograms of anti-FGFR2IIIb formulations for assessing charge variant profiles are shown. [Figure 9] Figure 1 shows the effect of buffer pH (in citrate or phosphate NaCl buffers) on acidic variant formation as determined by WCX-HPLC over the pH range 4-8 and over storage periods of 0-2 months at 25°C. There was a relatively large % increase in acidic variants in the pH 8 formulation, with the pH 7 formulation showing the next largest % increase. [Figure 10]Figure 1 shows the effect of buffer pH (in citrate or phosphate NaCl buffers) on basic variant formation as determined by WCX-HPLC over the pH range 4-8 and over storage periods of 0-2 months at 25°C. There was a relatively large % decrease in basic variants in the pH 8 formulation, with the pH 7 formulation showing the next largest % decrease. The smallest % change was observed at pH 4. [Figure 11] Figure 1 shows the effect of buffer pH (in citrate or phosphate NaCl buffers) on the size of the main peak in WCX-HPLC chromatograms ranging from pH 4 to 8 and over a storage period of 0 to 2 months at 25° C. The greatest % change in the main peak was observed in pH 4 and pH 8 formulations, and the least % change in pH 5, pH 6 and pH 7 formulations. [Figure 12] 1 shows the appearance of solutions of seven anti-FGFR2IIIb formulations shown in Table 8 below after shaking at 500 rpm for 72 hours. [Figure 13] Figure 1 shows the effect of excipients on anti-FGFR2IIIb stability over five freeze-thaw cycles at 70°C to ambient temperature. Formulations are as provided in Table 8. Only the histidine / NaCl formulation (number 4 in Table 8) appeared to show increased aggregates over multiple freeze-thaw cycles. [Figure 14] FIG. 1 shows the effect of buffers and bulking agents on anti-FGFR2IIIb aggregate formation over one month at 40° C. as determined by SE-HPLC. [Figure 15] FIG. 1 shows the effect of buffers and bulking agents on anti-FGFR2IIIb aggregate formation over 3 months at 40° C. as determined by SE-HPLC. [Figure 16] FIG. 1 shows the effect of buffers and bulking agents on anti-FGFR2IIIb acidic variant formation over 3 months at 25° C. as determined by WCX-HPLC. [Figure 17] 1 shows the effect of buffers and bulking agents on the size of the main WCX-HPLC peak over 3 months at 25° C. [Figure 18]1 shows the effect of buffers and bulking agents on the size of the main WCX-HPLC peak over 3 months at 25° C. [Figure 19] FIG. 1 shows the effect of mechanical stress over 72 h on histidine / arginine or histidine / sucrose anti-FGFR2IIIb antibody formulations at pH 5.5-6.5 as determined by analysis of aggregates by SE-HPLC. [Figure 20] FIG. 1 shows the effect of mechanical stress over 72 h on histidine / arginine or histidine / sucrose anti-FGFR2IIIb antibody formulations at pH 5.5-6.5 as determined by analysis of clips (fragments) by SE-HPLC. [Figure 21] 13 shows the effect of five freeze-thaw cycles from -70°C to ambient temperature on histidine / arginine or histidine / sucrose anti-FGFR2IIIb antibody formulations at pH 5.5-6.5 as determined by analysis of aggregates by SE-HPLC. [Figure 22] 1 shows the effect of five freeze-thaw cycles from -70°C to ambient temperature on histidine / arginine or histidine / sucrose anti-FGFR2IIIb antibody formulations at pH 5.5-6.5 as determined by analysis of clips (fragments) by SE-HPLC. [Figure 23] 1 shows the effect of pH between pH 5.5-6.5 on aggregate formation in a histidine / arginine formulation containing 20 mM histidine, 150 mM arginine, and 0.01% polysorbate 20. A shows aggregate formation at pH 5.5-6.0, showing that the % aggregates generally remained below 3% after 1 month at 40° C. B shows aggregate formation at pH 5.5-6.0, showing that the % aggregates generally remained below 2.5% after 3 months at 25° C. C shows aggregate formation at all pHs, showing that the % aggregates generally remained below about 1.5% and 2% after 6 months at 5° C. [Figure 24]1 shows the effect of pH between pH 5.5 and 6.5 on clips (fragments) in a histidine / arginine formulation containing 20 mM histidine, 150 mM arginine, and 0.01% polysorbate 20. A shows clip (fragment) formation at all pHs, showing that after 1 month at 40° C., the % clips generally remained about 1% or less than 1% at pH 6. B shows clip formation at all pHs, showing that after 3 months at 25° C., the % clips generally remained less than 1.5%. C shows clip formation at all pHs, showing that after 6 months at 5° C., the % clips generally remained less than 1% and in formulations at pH 5.7 or 6.0 generally remained less than 0.5%. [Diagram 25] FIG. 25 shows the effect of pH between pH 5.5-6.5 on aggregate formation in a histidine / sucrose formulation containing 20 mM histidine, 270 mM sucrose, and 0.01% polysorbate 20. FIG. 25A shows aggregate formation at pH 5.5-6.0, showing that the % aggregate generally remained below 4% after 1 month at 40° C., at pH 6.0 and 6.3 it was below 3%, and at pH 5.5 and 5.7 it was about 2.5%. B shows aggregate formation at pH 5.5-5.7, showing that the % aggregate generally remained below 2.0% after 3 months at 25° C., and at pH 6.0 it was below 2.5%. C shows aggregate formation at all pHs, showing that the % aggregate generally remained below about 1.5% and 2% after 6 months at 5° C. [Figure 26]1 shows the effect of pH between pH 5.5 and 6.5 on clips (fragments) in histidine / sucrose formulations containing 20 mM histidine, 270 mM sucrose, and 0.01% polysorbate 20. A shows clip (fragment) formation at all pHs, showing that the % clips remained generally below 2% after 1 month at 40° C. or below 1.5% at pH 5.5-6.3. B shows clip formation at all pHs, showing that the % clips remained generally below 3.5% after 3 months at 25° C., but that clip formation was pH dependent, with the lowest percentages at 1, 2, and 3 months for the pH 5.5 formulation, and the highest percentages every month for the pH 6.3 formulation. C shows clip formation at pH 6.0 or below, showing that the % clips remained generally below 1% after 6 months at 5° C. [Figure 27] 1 shows the effect of pH between pH 5.5-6.5 on acidic variant formation in histidine / sucrose formulations containing 20 mM histidine, 270 mM sucrose, and 0.01% polysorbate 20. A shows acidic variant formation at pH 5.5-6.0, showing that the % acidic variant generally remained below 40% after 1 month at 40° C. B shows acidic variant formation at pH 5.5-6.0, showing that the % acidic variant generally remained below 30.0% after 3 months at 25° C. C shows acidic variant formation at all pHs, showing that the % acidic variant generally remained below 25% after 6 months at 5° C. [Figure 28] 1 shows the effect of pH between pH 5.5-6.5 on acidic variant formation in histidine / arginine formulations containing 20 mM histidine, 150 mM arginine, and 0.01% polysorbate 20. A shows acidic variant formation at pH 5.5-6.0, showing that the % acidic variant generally remained below 40% after 1 month at 40° C. B shows acidic variant formation at pH 5.5-6.0, showing that the % acidic variant generally remained below 30.0% after 3 months at 25° C. C shows acidic variant formation at all pHs, showing that the % acidic variant generally remained below 25% after 6 months at 5° C. [Figure 29] 1 shows the effect of pH between pH 5.5-6.5 on basic variant formation in a histidine / sucrose formulation containing 20 mM histidine, 270 mM sucrose, and 0.01% polysorbate 20. A shows basic variant formation at pH 5.5-6.0, showing that the % basic variant remained mostly between 10%-20% after 1 month at 40° C. B shows basic variant formation at pH 5.5-6.0, showing that the % basic variant remained mostly between 20%-30% after 3 months at 25° C. C shows basic variant formation at all pHs, showing that the % basic variant remained mostly between 25%-30% after 6 months at 5° C. [Diagram 30] FIG. 30 shows the effect of pH between 5.5 and 6.5 on basic variant formation in a histidine / arginine formulation containing 20 mM histidine, 150 mM arginine, and 0.01% polysorbate 20. FIG. 30A shows basic variant formation at pH 5.5-6.0, showing that after 1 month at 40° C., the % basic variant remained mostly between 10%-20%. FIG. 30B shows basic variant formation at pH 5.5-6.0, showing that after 3 months at 25° C., the % basic variant remained generally between 15%-25%. FIG. 30C shows basic variant formation at all pHs, showing that after 6 months at 5° C., the % basic variant remained generally between 25%-30%.

[0018] For further details of the above figures, please see the Examples section herein. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0019] The section headings used herein are for organizational purposes only and should not be construed as limiting the subject matter described. All references cited herein, including patent applications and publications, are hereby incorporated by reference in their entirety for any purpose.

[0020] definition Unless otherwise defined, scientific and technical terms used in connection with the present invention shall have the meanings commonly understood by those of ordinary skill in the art. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular.

[0021] In this application, the use of "or" means "and / or" unless specifically stated otherwise. In the context of multiple dependent claims, the use of "or" refers to two or more preceding independent or dependent claims, in the alternative only. Also, terms such as "element" or "component" encompass both elements and components comprising a single unit, and elements and components comprising two or more subunits, unless specifically stated otherwise.

[0022] As described herein, any concentration range, percentage range, ratio range, or integer range, unless otherwise indicated, should be understood to include any integer value within the recited range, and fractions thereof (e.g., tenths and hundredths of integers), where appropriate.

[0023] Units, prefixes, and symbols are expressed in the form accepted by the Systeme International de Unites (SI). Numerical ranges are inclusive of the numbers defining the range. Measurements are understood to be approximate, taking into account significant orders of magnitude and errors associated with the measurements.

[0024] The headings provided herein are to be had by reference to the specification as a whole and are not limitations of the various aspects of the disclosure, and therefore the terms defined immediately below are more fully defined by reference to the specification as a whole.

[0025] As utilized in accordance with the present disclosure, the following terms, unless otherwise indicated, shall be understood to have the following meanings:

[0026] "Administering" refers to physically introducing a composition containing a therapeutic agent into a subject using any of a variety of methods and delivery systems known to those skilled in the art. Common routes of administration of antibodies include, for example, intravenous (IV), intramuscular, subcutaneous (SC), intraperitoneal, spinal, or other parenteral routes of administration by injection or infusion. As used herein, the phrase "parenteral administration" refers to a mode of administration other than enteral and topical administration, usually by injection, including, but not limited to, intravenous, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, intrasternal injection and infusion, and in vivo electroporation. Non-parenteral routes include, for example, topical, epidermal, or mucosal routes of administration, such as oral, intranasal, vaginal, rectal, sublingual, or topical. Administration can also be, for example, once, multiple times, and / or over one or more extended periods of time.

[0027] As used herein, a "pharmaceutical formulation" or "therapeutic formulation" refers to a composition containing a therapeutic agent, such as a suitable antibody, for pharmaceutical use, such as administration to a patient, either directly or after reconstitution, dilution, mixing, or dissolution with at least one additional composition, or after thawing from a frozen state. As used herein, a "ready-to-use" formulation refers to a formulation that can be administered directly to a patient and therefore does not need to be diluted, reconstituted, thawed from a frozen state, dissolved in solution, or mixed with other ingredients prior to administration.

[0028] As used herein, "subcutaneous administration" refers to administering a therapeutic agent under the skin. In some cases, subcutaneous administration (abbreviated SC or SubQ) can be performed using a needle connected to a syringe or pump, or using a special type of injector device (e.g., EpiPen®, etc.). In some cases, SC administration can release a therapeutic agent into the tissue between the skin layer and the muscle layer, and in some cases into the muscle layer.

[0029] As used herein, an "injectable" formulation is a formulation suitable for administration by SC injection.

[0030] As used herein, "intravenous administration" refers to administering a therapeutic agent directly into a vein of a subject. Intravenous administration can be by "intravenous infusion." Infusion refers to the use of gravity-supplied pressure to deliver a therapeutic agent to a patient.

[0031] The formulations herein may be contained in a "vial." A "vial" herein refers to any small container suitable for holding a liquid pharmaceutical formulation. In some cases, the "vial" may be connected to a syringe or pump, or the "vial" may be part of an administration device, for example, for injecting the contents of the vial into a patient. In some embodiments, the formulations may be contained in a "single-use vial." A "single-use vial" refers to a "vial" in which the entire vial or the remaining contents of the vial are discarded after the contents are administered.

[0032] The term "lyophilized" when applied to a formulation, material, or composition, refers to a formulation, material, or composition that has been lyophilized.

[0033] As used herein, the term "liquid formulation" refers to a formulation that is in a liquid state, including, for example, a solution or a liquid emulsion.

[0034] As used herein, a "buffer" is a substance that, when added to a solution or formulation, resists significant changes in pH upon dilution or addition of an acidic or basic component. Exemplary buffers herein include histidine, citrate, and phosphate buffers.

[0035] As referred to herein, an "isotonic" formulation is one that has approximately the same osmotic pressure as the human body fluid into which the formulation is introduced or that is present in the area of ​​administration, such as human plasma or human lymph.

[0036] "Aggregation" or "protein aggregation" in a formulation as used herein refers to the aggregation of polypeptide molecules in the formulation. The amount of polypeptide in a formulation that is in an aggregated state, such as a dimer or multimer, can be expressed, for example, as a percentage of the total polypeptide content of the formulation. Aggregation can be detected, for example, by size exclusion chromatography or other techniques that separate proteins in a solution based on size or molecular weight. In such techniques, the amount of "aggregates" can be equal to the amount of polypeptide in the formulation that has at least twice the molecular weight of the major polypeptide species of the formulation (e.g., anti-FGFR2-IIIb antibody). A species that has twice the molecular weight of the major polypeptide species of the formulation can be considered as a "dimer" herein, while a detectable species that has a molecular weight greater than that of a dimer can be referred to as a "higher molecular weight aggregate" or "multimer" or similar term.

[0037] When referring to the storage of a formulation herein, the phrase "in the dark" means that the formulation is stored so that it is protected from exposure to ambient light and ultraviolet light.For example, the formulation may be stored in a dark room or space, or in a package that protects the formulation from such light, or in a vial that has walls made of or is covered with a material that protects the contents from such light.

[0038] In some embodiments herein, the pharmaceutical formulations are "free" of one or more types of excipients or ingredients. In this context, "free" means that the excluded ingredient is not present beyond trace levels, for example, due to contamination or impurities found in other intentionally added ingredients.

[0039] When referring to a mixture of components of a formulation herein, the term "consisting essentially of" indicates that components other than those explicitly listed may be present, but that such components are found only in trace or otherwise sufficiently low amounts so as not to alter the fundamental characteristics of the formulation, including protein concentration, viscosity, thermal stability, osmolality, and pH.

[0040] The terms "polypeptide" and "protein" are used interchangeably to refer to a polymer of amino acid residues and are not limited to a minimum length. Such polymers of amino acid residues may contain natural or non-natural amino acid residues and include, but are not limited to, peptides, oligopeptides, dimers, trimers, and multimers of amino acid residues. Both full-length proteins and fragments thereof are encompassed by the definition. These terms also include post-expression modifications of the polypeptide, such as glycosylation, sialylation, acetylation, phosphorylation, and the like. Furthermore, for purposes of the present invention, "polypeptide" refers to a protein that includes modifications such as deletions, additions, and substitutions (generally conservative in nature) to the native sequence, so long as the protein maintains the desired activity. These modifications may be deliberate, via site-directed mutagenesis, or may be accidental, for example, via mutations of the host that produces the protein or errors resulting from PCR amplification.

[0041] "FGFR2" refers to human fibroblast growth factor receptor 2, including any of its alternative splice forms, such as IIIa, IIIb and IIIc splice forms, unless a non-human FGFR2 is explicitly indicated (e.g., "mouse FGFR2"). The term FGFR2 encompasses wild-type FGFR2 and naturally occurring mutant forms, such as FGFR2-activated mutant forms, e.g., FGFR2-S252W, found in some cancer cells. "FGFR2-IIIb" or "FGFR2b" are used interchangeably to refer to the fibroblast growth factor receptor 2 IIIb splice form. An exemplary human FGFR2-IIIb is set forth in GenBank Accession No. NP_075259.4, dated July 7, 2013. A non-limiting exemplary mature human FGFR2-IIIb amino acid sequence is set forth in SEQ ID NO:1. "FGFR2-IIIc" or "FGFR2c" are used interchangeably to refer to the fibroblast growth factor receptor 2 IIIc splice form. An exemplary human FGFR2-IIIc is set forth in GenBank Accession No. NP_000132.3, dated July 7, 2013. A non-limiting exemplary mature FGFR2-IIIc amino acid sequence is set forth in SEQ ID NO:13.

[0042] With respect to anti-FGFR2-IIIb antibodies, the term "blocking binding" or "inhibiting binding" of a ligand refers to the ability to inhibit the interaction between FGFR2-IIIb and an FGFR2 ligand, such as FGF1 or FGF2. Such inhibition can occur through any mechanism, including direct interference with ligand binding, for example, due to overlapping binding sites on FGFR2-IIIb and / or antibody-induced conformational changes in FGFR2-IIIb that alter ligand affinity.

[0043] "Affinity" or "binding affinity" refers to the strength of the sum of non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). In some embodiments, "binding affinity" refers to the intrinsic binding affinity that reflects a 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen). The affinity of a molecule X for a partner Y is generally determined by the dissociation constant (K d )

[0044] As used herein, the term "antibody" refers to a molecule that includes at least the hypervariable regions (HVRs) H1, H2, and H3 of a heavy chain and L1, L2, and L3 of a light chain, which molecule is capable of binding to an antigen. The term antibody includes full-length heavy and light chains, as well as Fv, single chain Fv (scFv), Fab, Fab', and (Fab') 2 Antibodies include, but are not limited to, antibodies that contain fragments capable of binding to antigens such as (also referred to herein as "antigen-binding fragments"). The term antibody also includes, but is not limited to, chimeric antibodies, humanized antibodies, human antibodies, and antibodies of various species such as mouse, human, cynomolgus monkey, etc. It also includes antibodies that are conjugated to other molecules, such as small molecule drugs, bispecific antibodies, and multispecific antibodies.

[0045] The term "heavy chain variable region" refers to a region comprising heavy chain HVR1, framework (FR) 2, HVR2, FR3, and HVR3. In some embodiments, the heavy chain variable region also comprises at least a portion of FR1 and / or at least a portion of FR4.

[0046] The term "heavy chain constant region" refers to a region of at least three heavy chain constant domains. H 1. C H 2, and C H The heavy chain constant region refers to a region that includes γ, δ, and α. Non-limiting exemplary heavy chain constant regions also include ε and μ. Each heavy constant region corresponds to an antibody isotype. For example, an antibody that includes a γ constant region is an IgG antibody, an antibody that includes a δ constant region is an IgD antibody, and an antibody that includes an α constant region is an IgA antibody. Furthermore, an antibody that includes a μ constant region is an IgM antibody, and an antibody that includes an ε constant region is an IgE antibody. Certain isotypes can be further subdivided into subclasses. For example, IgG antibodies include, but are not limited to, IgG1 (γ 1 IgG2(γ 2 (including constant region), IgG3(γ 3 IgG4 (including the constant region) and IgG4 (γ 4 Examples of IgA antibodies include, but are not limited to, IgA1 (α 1 IgA2 (including the constant region) and IgA2 (α 2 IgM antibodies include, but are not limited to, IgM1 and IgM2.

[0047] The term "heavy chain" refers to a polypeptide comprising at least a heavy chain variable region, with or without a leader sequence. In some embodiments, a heavy chain includes at least a portion of a heavy chain constant region. The term "full-length heavy chain" refers to a polypeptide comprising a heavy chain variable region and a heavy chain constant region, with or without a leader sequence.

[0048] The term "light chain variable region" refers to a region comprising light chain HVR1, framework (FR) 2, HVR2, FR3, and HVR3. In some embodiments, the light chain variable region also comprises FR1 and / or FR4.

[0049] The term "light chain constant region" refers to a light chain constant domain C L Non-limiting exemplary light chain constant regions include lambda and kappa.

[0050] The term "light chain" refers to a polypeptide comprising at least a light chain variable region, with or without a leader sequence. In some embodiments, a light chain comprises at least a portion of a light chain constant region. The term "full light chain" refers to a polypeptide comprising a light chain variable region and a light chain constant region, with or without a leader sequence.

[0051] The term "hypervariable region" or "HVR" refers to each of the regions of an antibody variable domain that are hypervariable in sequence and / or form structurally defined loops ("hypervariable loops"). Generally, a naturally occurring four-chain antibody contains six HVRs, H 3 in (H1, H2, H3), V LHVRs generally comprise amino acid residues from the hypervariable loops and / or from the "complementarity determining regions" (CDRs), the latter of which have the highest sequence variability and / or are involved in antigen recognition. Exemplary hypervariable loops occur at amino acid residues 26-32 (L1), 50-52 (L2), 91-96 (L3), 26-32 (H1), 53-55 (H2), and 96-101 (H3) (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987).). Exemplary CDRs (CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2, and CDR-H3) occur at amino acid residues 24-34 of L1, amino acid residues 50-56 of L2, amino acid residues 89-97 of L3, amino acid residues 31-35B of H1, amino acid residues 50-65 of H2, and amino acid residues 95-102 of H3 (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD (1991)). The terms hypervariable region (HVR) and complementarity determining region (CDR) both refer to the portions of the variable regions that form the antigen binding region.

[0052] As used herein, a "chimeric antibody" refers to an antibody that comprises at least one variable region derived from a first species (mouse, rat, cynomolgus monkey, etc.) and at least one constant region derived from a second species (human, cynomolgus monkey, etc.). In some embodiments, a chimeric antibody comprises at least one mouse variable region and at least one human constant region. In some embodiments, a chimeric antibody comprises at least one cynomolgus monkey variable region and at least one human constant region. In some embodiments, a chimeric antibody comprises at least one rat variable region and at least one mouse constant region. In some embodiments, all of the variable regions of a chimeric antibody are derived from a first species and all of the constant regions of a chimeric antibody are derived from a second species.

[0053] As used herein, "humanized antibody" refers to an antibody in which at least one amino acid in a framework region of a non-human variable region is replaced with the corresponding amino acid from a human variable region. In some embodiments, a humanized antibody comprises at least one human constant region or a fragment thereof. In some embodiments, a humanized antibody is an antibody that is a Fab, scFv, (Fab') 2 etc.

[0054] As used herein, "CDR-grafted antibody" or "HVR-grafted antibody" refers to a humanized antibody in which the complementarity determining regions (CDRs) or hypervariable regions (HVRs) of a first (non-human) species have been grafted onto the framework regions (FRs) of a second (human) species.

[0055] As used herein, "human antibody" refers to antibodies produced in humans, antibodies produced in non-human animals that contain human immunoglobulin genes, such as the XenoMouse®, and antibodies selected using in vitro methods, such as phage display, where the antibody repertoire is based on human immunoglobulin sequences.

[0056] "Afucosylated" or "fucose-devoid" antibodies refer to IgG1 or IgG3 isotype antibodies lacking fucose in their constant region glycosylation. Glycosylation of human IgG1 or IgG3 occurs as a core fucosylated biantennary complex oligosaccharide glycosylation terminated with up to 2 Gal residues at Asn297 (N297). In some embodiments, afucosylated antibodies lack fucose at Asn297. These structures are designated as G0, G1 (α1,6 or α1,3) or G2 glycan residues depending on the amount of terminal Gal residues. See, e.g., Raju, TS, BioProcess Int. 1:44-53 (2003). For CHO-type glycosylation of antibody Fc, see, e.g., Routier, FH, Glycoconjugate J. 14:201-207 (1997). Within a population of antibodies, an antibody is considered to be afucosylated if less than 5% of the antibodies in the population contain fucose at Asn297.

[0057] "Effector function" refers to the biological activities attributable to the Fc region of an antibody, which vary depending on the antibody isotype. Examples of antibody effector functions include Clq binding and complement-dependent cytotoxicity (CDC), Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, down-regulation of cell surface receptors (e.g., B cell receptor), and B cell activation.

[0058] "Antibody-dependent cell-mediated cytotoxicity" or "ADCC" refers to a form of cytotoxicity in which secreted Ig bound to Fc receptors (FcR) present on certain cytotoxic cells (e.g., NK cells, neutrophils, and macrophages) enables these cytotoxic effector cells to specifically bind to antigen-bearing target cells and subsequently kill the target cells with cytotoxins. NK cells, the primary cells for mediating ADCC, express only FcγRIII, whereas monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol 9:457-92 (1991). To assess the ADCC activity of a molecule of interest, an in vitro ADCC assay may be performed, such as those described in U.S. Pat. Nos. 5,500,362 or 5,821,337 or U.S. Pat. No. 6,737,056 (Presta). Effector cells useful for such assays include PBMCs and NK cells. Alternatively, or in addition, the ADCC activity of a molecule of interest may be assessed in vivo, for example in an animal model, such as those disclosed in Clynes et al. Proc. Natl. Acad. Sci. (USA) 95:652-656 (1998). Additional antibodies with altered Fc region amino acid sequences to increase or decrease ADCC activity are described, for example, in U.S. Pat. Nos. 7,923,538 and 7,994,290.

[0059] An antibody with "improved ADCC activity" refers to an antibody that is more effective at mediating ADCC in vitro or in vivo compared to a parent antibody (wherein the antibody and parent antibody differ in at least one structural aspect) when essentially the same amount of such antibody and parent antibody are used in the assay. In some embodiments, the antibody and parent antibody have the same amino acid sequence, but the antibody is afucosylated while the parent antibody is fucosylated. In some embodiments, ADCC activity can be determined using the in vitro ADCC assay disclosed in US Publication No. 2015 / 0050273A1, although other assays or methods for determining ADCC activity, e.g., animal models, are contemplated. In other embodiments, an antibody with improved ADCC activity also has improved affinity for FcγRIIIA. In some embodiments, an antibody with improved ADCC activity has improved affinity for FcγRIIIA(V158). In certain embodiments, an antibody with improved ADCC activity has improved affinity for FcγRIIIA(F158).

[0060] "Improved affinity for FcγRIIIA" refers to an antibody that has a higher affinity for FcγRIIIA (sometimes referred to as CD16a) than the parent antibody, where the antibody and the parent antibody differ in at least one structural aspect. In some embodiments, the antibody and the parent antibody have the same amino acid sequence, but the parent antibody is fucosylated while the antibody is afucosylated. Any suitable method for determining affinity for FcγRIIIA may be used. In some embodiments, affinity for FcγRIIIA is determined by the methods described in US Publication No. 2015 / 0050273-A1. In other embodiments, antibodies with improved affinity for FcγRIIIA also have improved ADCC activity. In certain embodiments, antibodies with improved affinity for FcγRIIIA have improved affinity for FcγRIIIA(V158). In some embodiments, antibodies with improved affinity for FcγRIIIA have improved affinity for FcγRIIIA(F158).

[0061] The term "leader sequence" refers to a sequence of amino acid residues located at the N-terminus of a polypeptide that facilitates secretion of the polypeptide from a mammalian cell. The leader sequence may be cleaved upon export of the polypeptide from the mammalian cell to form the mature protein. Leader sequences may be natural or synthetic, and they may be heterologous or homologous to the protein to which they bind. Non-limiting exemplary leader sequences include leader sequences from heterologous proteins. In some embodiments, the antibody lacks a leader sequence. In other embodiments, the antibody comprises at least one leader sequence, which may be selected from a natural antibody leader sequence and a heterologous leader sequence.

[0062] As used herein, the term "isolated" refers to a molecule that is separated from at least some of the components that it typically finds in nature. For example, a polypeptide is referred to as "isolated" when it is separated from at least some of the components of the cell in which it is produced. If the polypeptide is secreted by the cell after expression, physically separating the supernatant containing the polypeptide from the cell that produced it is considered to "isolate" the polypeptide. Similarly, a polynucleotide is referred to as "isolated" when it is not part of a larger polynucleotide that it typically finds in nature (e.g., genomic DNA or mitochondrial DNA in the case of a DNA polynucleotide), or when it is separated from at least some of the components of the cell in which it is produced, for example, in the case of an RNA polynucleotide. Thus, a DNA polynucleotide contained in a vector in a host cell can be referred to as "isolated" as long as the polynucleotide is not naturally found in the vector.

[0063] The terms "subject" and "patient" are used interchangeably herein to refer to humans. In some embodiments, methods of treating other mammals are also provided, including, but not limited to, rodents, monkeys, cats, dogs, horses, cows, pigs, sheep, goats, mammalian laboratory animals, mammalian farm animals, mammalian sports animals, and mammalian pets.

[0064] The term "cancer" is used herein to refer to a group of cells that exhibits abnormally high levels of proliferation and growth. Cancers can be benign (also referred to as benign tumors), pre-malignant, or malignant. Cancer cells can be solid cancer cells or leukemic cancer cells. The term "cancer growth" is used herein to refer to proliferation or growth by one or more cells that comprise the cancer, resulting in a corresponding increase in size or extent of the cancer.

[0065] Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and leukemia. More specific non-limiting examples of such cancer include squamous cell carcinoma, small cell lung cancer, pituitary cancer, esophageal cancer, astrocytoma, soft tissue sarcoma, non-small cell lung cancer (including squamous cell non-small cell lung cancer), adenocarcinoma of the lung, squamous cell carcinoma of the lung, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatocellular carcinoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, renal cell carcinoma, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, liver cancer, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, gastric cancer, melanoma, and various types of head and neck cancer (including squamous cell carcinoma of head and neck cancer).

[0066] As used herein, "treatment" refers to therapeutic treatment, for example, to cure a condition or disorder, to reduce the severity or slow the progression of a condition or disorder, or to inhibit the recurrence of a condition or disorder. In certain embodiments, the term "treatment" encompasses any administration or application of a therapeutic agent for a disease in a patient, including inhibiting or slowing the progression of a disease or disease, for example, partially or completely relieving a disease by causing degeneration or by restoring or repairing a lost, missing, or defective function, stimulating an inefficient process, or reducing the severity of a stable disease. The term "treatment" also includes reducing the severity of any phenotypic characteristic and / or reducing the incidence, extent, or likelihood of the characteristic. Those in need of treatment include those already with the disorder, as well as those at risk of recurrence of the disorder or those in need of preventing or slowing down the recurrence of the disorder.

[0067] The term "effective amount" or "therapeutically effective amount" refers to an amount of drug effective to treat a disease or disorder in a subject. In certain embodiments, an effective amount refers to an amount effective at the dosage and duration necessary to achieve a desired therapeutic result. The therapeutically effective amount of the anti-FGFR2 antibody of the present invention may vary depending on factors such as the disease state, age, sex, and weight of the individual, and the ability of the one or more antibodies to elicit a desired response in the individual. A therapeutically effective amount encompasses an amount in which any toxic or harmful effects of the one or more antibodies are outweighed by the therapeutically beneficial effects. In some embodiments, the term "effective amount" refers to an amount of an antibody effective to treat cancer.

[0068] Additional definitions are provided in the following sections.

[0069] Anti-FGFR2 antibody In any of the formulations described herein that include an anti-FGFR2 antibody, the anti-FGFR2 antibody can be a monoclonal antibody, a genetically engineered antibody, a humanized antibody, a chimeric antibody, or a human antibody. In any of the compositions or methods described herein, the anti-FGFR2 antibody can be selected from Fab, Fv, scFv, Fab', and (Fab')2. In any of the compositions or formulations described herein, the anti-FGFR2 antibody can be selected from IgA, IgG, and IgD. In any of the compositions or formulations described herein, the anti-FGFR2 antibody can be IgG. In any of the methods described herein, the antibody can be IgG1 or IgG3.

[0070] Exemplary anti-FGFR2 antibodies include antibodies that bind to FGFR2-IIIb. In some embodiments, anti-FGFR2-IIIb antibodies bind to FGFR2-IIIc with lower affinity than they bind to FGFR2-IIIb. In other embodiments, anti-FGFR2-IIIb antibodies do not detectably bind to FGFR2-IIIc.

[0071] An exemplary anti-FGFR2-IIIb antibody for use in the embodiments herein is the HuGAL-FR21 antibody described in U.S. Patent No. 8,101,723B2, issued on January 24, 2012, which is specifically incorporated herein by reference. Figures 13 and 14 of U.S. Patent No. 8,101,723B2 show the amino acid sequences of the variable region and full-length mature antibody chain of HuGAL-FR21, which are specifically incorporated herein by reference. The heavy chain variable region sequence of antibody HuGAL-FR21 is underlined in Figure 13 of U.S. Patent No. 8,101,723B2, which is specifically incorporated herein by reference. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297. Additional antibodies that may be used in embodiments herein include those described in U.S. Patent Publication No. 2015-0050273-A1, which describes certain afucosylated FGFR2-IIIb antibodies and is incorporated herein by reference.

[0072] In some embodiments, the anti-FGFR2-IIIb antibody comprises at least one, two, three, four, five, or six hypervariable regions (HVRs, e.g., CDRs) selected from: (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:8; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:9; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:10; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:11. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0073] In some embodiments, the anti-FGFR2-IIIb antibody comprises a heavy chain variable region and a light chain variable region. In some embodiments, the anti-FGFR2-IIIb antibody comprises at least one heavy chain comprising a heavy chain variable region and at least a portion of a heavy chain constant region, and at least one light chain comprising a light chain variable region and at least a portion of a light chain constant region. In some embodiments, the anti-FGFR2-IIIb antibody comprises two heavy chains (each heavy chain comprises at least a portion of a heavy chain variable region and a heavy chain constant region), and two light chains (each light chain comprises at least a portion of a light chain variable region and a light chain constant region). In some embodiments, the anti-FGFR2-IIIb antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO:4, and a light chain variable region comprising the amino acid sequence of SEQ ID NO:5. In some embodiments, the anti-FGFR2-IIIb antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:2, and a light chain comprising the amino acid sequence of SEQ ID NO:3. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody that lacks fucose at Asn297.

[0074] In some embodiments, the anti-FGFR2-IIIb antibody comprises six HVRs, including (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6, (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7, (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:8, (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:9, (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:10, and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:11. In some embodiments, the anti-FGFR2-IIIb antibody comprises the six HVRs described above and binds to FGFR2-IIIb. In some embodiments, the anti-FGFR-IIIb antibody does not bind to FGFR2-IIIc. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0075] In one aspect, the anti-FGFR2-IIIb antibody competes with an anti-FGFR2-IIIb antibody comprising six HVRs, including: (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:8; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:9; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:10; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:11. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0076] In some embodiments, the anti-FGFR2-IIIb antibody comprises at least one, at least two, or all three VH HVR sequences selected from (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6, (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7, and (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:8. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0077] In some embodiments, the anti-FGFR2-IIIb antibody comprises at least one, at least two, or all three VL HVR sequences selected from (a) HVR-L1 comprising the amino acid sequence of SEQ ID NO:9, (b) HVR-L2 comprising the amino acid sequence of SEQ ID NO:10, and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO:11. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0078] In some embodiments, the anti-FGFR2-IIIb antibody comprises: (a) a VH domain comprising at least one, at least two, or all three VH HVR sequences selected from (i) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6, (ii) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7, and (iii) HVR-H3 comprising the amino acid sequence selected from SEQ ID NO:8; and (b) a VL domain comprising at least one, at least two, or all three VL HVR sequences selected from (i) HVR-L1 comprising the amino acid sequence of SEQ ID NO:9, (ii) HVR-L2 comprising the amino acid sequence of SEQ ID NO:10, and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO:11. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0079] In some embodiments, anti-FGFR2-IIIb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of SEQ ID NO: 4. In certain embodiments, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions or deletions compared to the reference sequence, but the anti-FGFR2-IIIb antibody comprising the sequence retains the ability to bind to FGFR2-IIIb. In certain embodiments, such FGFR2-IIIb antibody retains the ability to selectively bind to FGFR2-IIIb without detectably binding to FGFR2-IIIc. In certain embodiments, a total of 1-10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO:4. In certain embodiments, the substitutions, insertions, or deletions occur in the regions outside the HVRs (i.e., FRs). Optionally, the anti-FGFR2-IIIb antibody comprises a VH sequence of SEQ ID NO:5 (including post-translational modifications of that sequence). In certain embodiments, the VH comprises one, two, or three HVRs selected from (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6, (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7, and (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:8. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0080] In some embodiments, anti-FGFR2-IIIb antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence of SEQ ID NO:5. In certain embodiments, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity contains substitutions (e.g., conservative substitutions), insertions or deletions compared to the reference sequence, but the anti-FGFR2-IIIb antibody comprising the sequence retains the ability to bind to FGFR2-IIIb. In certain embodiments, anti-FGFR2-IIIb antibody retains the ability to selectively bind to FGFR2-IIIb without binding to FGFR2-IIIc. In certain embodiments, a total of 1-10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO:5. In certain embodiments, the substitutions, insertions, or deletions occur in the regions outside the HVRs (i.e., FRs). Optionally, the anti-FGFR2-IIIb antibody comprises a VL sequence of SEQ ID NO:4 (including post-translational modifications of that sequence). In certain embodiments, the VL comprises one, two, or three HVRs selected from (a) HVR-L1 comprising the amino acid sequence of SEQ ID NO:9, (b) HVR-L2 comprising the amino acid sequence of SEQ ID NO:10, and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO:11. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0081] In some embodiments, the anti-FGFR2-IIIb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO:4, and a light chain variable domain (VL) having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO:5. In certain embodiments, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, and a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-FGFR2-IIIb antibody comprising the sequence retains the ability to bind to FGFR2-IIIb. In certain embodiments, such an anti-FGFR2-IIIb antibody retains the ability to selectively bind to FGFR2-IIIb without binding to FGFR2-IIIc. In certain embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO:4. In certain embodiments, a total of 1-10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO:5. In certain embodiments, the substitutions, insertions, or deletions occur in the regions outside the HVRs (i.e., FRs). Optionally, the anti-FGFR2-IIIb antibody comprises a VH sequence of SEQ ID NO:4 and a VL sequence of SEQ ID NO:5, including post-translational modifications of one or both sequences. In certain embodiments, the VH comprises one, two, or three HVRs selected from (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6, (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7, and (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:8, and the VL comprises one, two, or three HVRs selected from (a) HVR-L1 comprising the amino acid sequence of SEQ ID NO:9, (b) HVR-L2 comprising the amino acid sequence of SEQ ID NO:10, and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO:11.In some embodiments, the antibody is afucosylated, hi some embodiments, the antibody is an IgG1 or IgG3 antibody that lacks fucose at Asn297.

[0082] In some embodiments, the anti-FGFR2-IIIb antibody comprises a VH as in any of the embodiments provided above, and a VL as in any of the embodiments provided above. In one embodiment, the antibody comprises the VH and VL sequences of SEQ ID NO: 4 and SEQ ID NO: 5, respectively, including post-translational modifications of those sequences. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0083] In some embodiments, the anti-FGFR2-IIIb antibody comprises a heavy chain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO:2. In certain embodiments, the heavy chain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions compared to the reference sequence, but the anti-FGFR2-IIIb antibody comprising the sequence retains the ability to bind to FGFR2-IIIb. In certain embodiments, such an anti-FGFR2-IIIb antibody retains the ability to selectively bind to FGFR2-IIIb without detectably binding to FGFR2-IIIc. In certain embodiments, a total of 1-10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO:2. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., FRs). Optionally, the anti-FGFR2-IIIb antibody comprises a VH sequence of SEQ ID NO:2 (including post-translational modifications of that sequence). In certain embodiments, the heavy chain comprises one, two, or three HVRs selected from (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6, (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7, and (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:8. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0084] In some embodiments, the anti-FGFR2-IIIb antibody comprises a light chain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 3. In certain embodiments, the light chain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions compared to the reference sequence, but the anti-FGFR2-IIIb antibody comprising the sequence retains the ability to bind to FGFR2-IIIb. In certain embodiments, such an anti-FGFR2-IIIb antibody retains the ability to selectively bind to FGFR2-IIIb without detectably binding to FGFR2-IIIc. In certain embodiments, a total of 1-10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 3. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside the HVRs (i.e., FRs). Optionally, the anti-FGFR2-IIIb antibody comprises a VL sequence of SEQ ID NO: 3 (including post-translational modifications of that sequence). In certain embodiments, the light chain comprises one, two, or three HVRs selected from (a) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 9, (b) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 10, and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 11. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0085] In some embodiments, the anti-FGFR2-IIIb antibody comprises a heavy chain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 2, and a light chain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 3. In certain embodiments, the heavy chain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions compared to the reference sequence, but the anti-FGFR2-IIIb antibody comprising the sequence retains the ability to bind to FGFR2-IIIb. In certain embodiments, such anti-FGFR2-IIIb antibodies retain the ability to selectively bind to FGFR2-IIIb without detectably binding to FGFR2-IIIc. In certain embodiments, a light chain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions compared to a reference sequence, but an anti-FGFR2-IIIb antibody comprising the sequence retains the ability to bind to FGFR2-IIIb. In certain embodiments, such anti-FGFR2-IIIb antibodies retain the ability to selectively bind to FGFR2-IIIb without detectably binding to FGFR2-IIIc. In certain embodiments, a total of 1-10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO:2. In certain embodiments, a total of 1-10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO:3. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside the HVR (i.e., FR). Optionally, the anti-FGFR2-IIIb antibody heavy chain comprises the VH sequence of SEQ ID NO: 2 (including post-translational modifications of that sequence), and the anti-FGFR2-IIIb antibody light chain comprises the VL sequence of SEQ ID NO: 3 (including post-translational modifications of that sequence).In certain embodiments, the heavy chain comprises one, two, or three HVRs selected from (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6, (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:7, and (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:8, and the light chain comprises one, two, or three HVRs selected from (a) HVR-L1 comprising the amino acid sequence of SEQ ID NO:9, (b) HVR-L2 comprising the amino acid sequence of SEQ ID NO:10, and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO:11. In some embodiments, the antibody is afucosylated. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0086] Further exemplary anti-FGFR2 antibodies are the GAL-FR22 and GAL-FR23 antibodies described in U.S. Patent No. 8,101,723B2, which is incorporated herein by reference. For example, the light and heavy chain variable regions of GAL-FR22 are provided as SEQ ID NOs: 7 and 8 in U.S. Patent No. 8,101,723B2, while the Kabat CDRs and light and heavy chain variable regions are also provided in FIG. 16 of that patent, which is incorporated herein by reference. Hybridomas producing GAL-FR21, GAL-FR22 and GAL-FR23 have been deposited at the American Type Culture Collection (PO Box 1549, Manassas VA, USA, 20108) under ATCC numbers 9586, 9587 and 9408 on November 6, 2008, November 6 and August 12, 2008, respectively. Thus, in some embodiments, the anti-FGFR2 antibody is an antibody that comprises the amino acid sequence of an antibody obtained from one of these three hybridoma lines.

[0087] The heavy and light chain variable regions of GAL-FR22 are also presented herein as SEQ ID NOs: 15 and 19, while the Kabat CDRs are presented herein as SEQ ID NOs: 16-19 and 20-22. Thus, in some embodiments, the anti-FGFR2-IIIb antibody heavy chain variable region comprises (i) HVR1 (CDR1) comprising the amino acid sequence of SEQ ID NO: 16, (ii) HVR2 comprising the amino acid sequence of SEQ ID NO: 17, and (iii) HVR3 comprising the amino acid sequence of SEQ ID NO: 18, and the light chain variable region comprises (iv) HVR1 comprising the amino acid sequence of SEQ ID NO: 20, (v) HVR2 comprising the amino acid sequence of SEQ ID NO: 21, and (vi) HVR3 comprising the amino acid sequence of SEQ ID NO: 22.

[0088] In some embodiments, the anti-FGFR2 antibody comprises an FGFR2-IIIb antibody, the heavy chain variable domain of which is at least 95%, e.g., at least 97%, at least 98%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 15, or comprises the amino acid sequence of SEQ ID NO: 15. In some embodiments, the anti-FGFR2 antibody comprises an FGFR2-IIIb antibody, the light chain variable domain of which is at least 95%, e.g., at least 97%, at least 98%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 19, or comprises the amino acid sequence of SEQ ID NO: 19. In some embodiments, the heavy chain variable domain of which is at least 95%, e.g., at least 97%, at least 98%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 15, or comprises the amino acid sequence of SEQ ID NO: 39, and the light chain variable domain of which is at least 95%, e.g., at least 97%, at least 98%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 19, or comprises the amino acid sequence of SEQ ID NO: 19. In some such embodiments, the anti-FGFR2-IIIb antibody heavy chain variable region also comprises (i) a HVR1 (CDR1) comprising the amino acid sequence of SEQ ID NO: 16, (ii) a HVR2 comprising the amino acid sequence of SEQ ID NO: 17, and (iii) a HVR3 comprising the amino acid sequence of SEQ ID NO: 18, and / or the light chain variable region also comprises (iv) a HVR1 comprising the amino acid sequence of SEQ ID NO: 20, (v) a HVR2 comprising the amino acid sequence of SEQ ID NO: 21, and (vi) a HVR3 comprising the amino acid sequence of SEQ ID NO: 22. In some embodiments, the antibody is an IgG1 or IgG3 antibody lacking fucose at Asn297.

[0089] Afucosylated anti-FGFR2 antibody In some embodiments, the anti-FGFR2 antibody described herein has a carbohydrate structure that lacks fucose attached (directly or indirectly) to the Fc region, i.e., the antibody is afucosylated. In some embodiments, the afucosylated antibody is an IgG1 or IgG3 antibody that lacks fucose at Asn297.

[0090] As used herein, an antibody is considered "afucosylated" if a plurality of such antibodies comprises at least 95% afucosylated antibody. The amount of fucose can be determined by calculating the average amount of fucose in the glycan at Asn297 compared to the sum of all glycan structures (e.g., complex, hybrid, and high mannose structures) attached to Asn297. Non-limiting exemplary methods for detection of fucose in antibodies include MALDI-TOF mass spectrometry (see, e.g., WO 2008 / 077546), HPLC measurement of released fluorescently labeled oligosaccharides (see, e.g., Schneider et al., "N-Glycan analysis of monoclonal antibodies and other glycoproteins using UHPLC with fluorescence detection," Agilent Technologies, Inc. (2012); Lines, J. Pharm. Biomed. Analysis, 14:601-608 (1996); Takahasi, J. Chrom., 720:217-225 (1996)), capillary electrophoresis measurement of released fluorescently labeled oligosaccharides (see, e.g., Ma et al., Anal. Chem., 71:5185-5192 (1999)), and HPLC with pulsed amperometric detection to measure monosaccharide composition (see, e.g., Hardy, et al., "N-Glycan analysis of monoclonal antibodies and other glycoproteins using UHPLC with fluorescence detection," Agilent Technologies, Inc. (2012); Lines, J. Pharm. Biomed. Analysis, 14:601-608 (1996); Takahasi, J. Chrom., 720:217-225 (1996)). al., Analytical Biochem., 170:54-62 (1988). In some embodiments, in the afucosylated antibody compositions herein, fucose is undetectable by one or more of these methods.

[0091] Asn297 refers to an asparagine residue located at about position 297 (EU numbering of Fc region residues) in the Fc region, although in a given antibody sequence, Asn297 may be located about ±3 amino acids upstream or downstream of position 297, i.e., between positions 294 and 300, due to minor sequence variations in antibodies. In the FGFR2-IIIb antibodies described herein, Asn297 is This is found in TIFF2024138284000001.tif7166 and is shown in bold and underlined in SEQ ID NO:2 of the sequence listing provided below.

[0092] Fucosylated variants may have improved ADCC function. See, e.g., U.S. Patent Publication Nos. 2003 / 0157108 (Presta, L.) and 2004 / 0093621 (Kyowa Hakko Kogyo Co., Ltd). Examples of publications related to "afucosylated" or "fucose-deficient" antibodies include US2003 / 0157108, WO2000 / 61739, WO2001 / 29246, US2003 / 0115614, US2002 / 0164328, US2004 / 0093621, US2004 / 0132140, US2004 / 0110704, US2004 / 0110282, US2004 / 0109865, WO2003 / 085119, WO2003 / 084570, WO2005 / 035586, WO2005 / 035778, WO2005 / 053742, WO2002 / 031140, Okazaki et al. J. Mol. Biol. 336: 1239-1249 (2004), and Yamane-Ohnuki et al. Biotech. Bioeng. 87: 614 (2004). Examples of cell lines capable of producing afucosylated antibodies include Lec13 CHO cells, which lack protein fucosylation (Ripka et al. Arch. Biochem. Biophys. 249:533-545 (1986); Presta, L, U.S. Patent Application No. US 2003 / 0157108 A1; and Adams et al., WO 2004 / 056312 A1, especially Example 11), as well as knockout cell lines, such as cell lines lacking a functional α-1,6-fucosyltransferase gene, FUT8, e.g. knockout CHO cells (e.g. Yamane-Ohnuki et al. Biotech. Bioeng. 87:614 (2004); Kanda, Y. et al., J. Immunol. 1999, 14:1311 (2004); al., Biotechnol. Bioeng., 94(4):680-688 (2006), and WO2003 / 085107).

[0093] The anti-FGFR2 antibody herein may also have biantennary oligosaccharides, for example, biantennary oligosaccharides attached to the Fc region of the antibody are biantennary by GlcNAc. Such antibodies may have reduced fucosylation and / or improved ADCC function. Examples of such antibodies are described, for example, in WO2003 / 011878 (Jean-Mairet et al.), U.S. Patent No. 6,602,684 (Umana et al.), and US2005 / 0123546 (Umana et al.). In some embodiments, the FGFR2 antibody has at least one galactose residue in the oligosaccharide attached to the Fc region. Such antibodies may have improved CDC function. Such antibodies are described, for example, in WO1997 / 30087 (Patel et al.), WO1998 / 58964 (Raju, S.), and WO1999 / 22764 (Raju, S.).

[0094] In certain embodiments of the present invention, the afucosylated FGFR2 antibody mediates ADCC in the presence of human effector cells more effectively than an antibody having the same amino acid sequence containing fucose. Generally, ADCC activity can be determined using the in vitro ADCC assay disclosed in US Publication No. 2015-0050273A1, although other assays or methods for determining ADCC activity are contemplated, such as animal models.

[0095] In some embodiments, the FGFR2 antibody comprises the heavy and light chain sequences of SEQ ID NOs: 2 and 3. In other embodiments, the antibody comprising the heavy and light chain sequences of SEQ ID NOs: 2 and 3 is afucosylated.

[0096] Pharmaceutical formulations of anti-FGFR2 antibodies The present disclosure provides pharmaceutical formulations of anti-FGFR2 antibodies suitable for administration to humans.Anti-FGFR2 antibodies can be any such antibodies disclosed herein.The amount and type of composition in the pharmaceutical formulation is selected to provide maximum stability to the antibody and thus maximum shelf life.

[0097] The pharmaceutical formulation may comprise, for example, an anti-FGFR2 antibody, a buffer selected from histidine, citrate, and phosphate, sucrose, and a surfactant. In some embodiments, the formulation consists essentially of an anti-FGFR2 antibody, a buffer selected from histidine, citrate, and phosphate, sucrose, and a surfactant. The pharmaceutical formulation may alternatively comprise, for example, an anti-FGFR2 antibody, a buffer selected from histidine, citrate, and phosphate, arginine, and a surfactant. In some embodiments, the formulation consists essentially of an anti-FGFR2 antibody, a buffer selected from histidine, citrate, and phosphate, arginine, and a surfactant. The pharmaceutical formulation may comprise, for example, an anti-FGFR2 antibody, histidine, sucrose, and a surfactant. In some embodiments, the formulation consists essentially of an anti-FGFR2 antibody, histidine, sucrose, and a surfactant. The pharmaceutical formulation may alternatively include, for example, an anti-FGFR2 antibody, histidine, arginine, and a surfactant. In some embodiments, the formulation consists essentially of an anti-FGFR2 antibody, histidine, arginine, and a surfactant. In some embodiments, the formulation is suitable for intravenous administration to a patient, for example, by infusion or injection. In other embodiments, the formulation is suitable for subcutaneous administration to a patient, for example, by injection. In certain embodiments, the formulation is isotonic with the body fluid in the area of ​​administration, such as human plasma or lymphatic fluid.

[0098] In some embodiments, the anti-FGFR2 antibodies herein have been found to be soluble in aqueous formulations at up to 180 mg / mL. In some embodiments, the formulation comprises 10-180 mg / mL of anti-FGFR2 antibody, or 10-30 mg / mL of anti-FGFR2 antibody, or, for example, 10-25 mg / mL, 20-30 mg / mL, 15-25 mg / mL, 10-15 mg / mL, 15-20 mg / mL, 20-25 mg / mL, 18-22 mg / mL, 10 mg / mL, 11 mg / mL, 12 mg / mL, 13 mg / mL, 14 mg / mL, 15 mg / mL, 16 mg / mL, 17 mg / mL, 18 mg / mL, 19 mg / mL, 20 mg / mL, 21 mg / mL, 22 mg / mL, 23 mg / mL, 24 mg / mL, 25 mg / mL, 26 mg / mL, 27 mg / mL, 28 mg / mL, 29 mg / mL, or 30 mg / mL of anti-FGFR2 antibody. In some embodiments, the formulation contains 20 mg / mL of antibody.

[0099] In some embodiments, the formulation comprises histidine, citrate, or phosphate as a buffer. In some such cases, the formulation buffer consists essentially of histidine. In some embodiments, the formulation comprises 5-40 mM, e.g., 10-40 mM, 10-30 mM, 15-25 mM, 10-20 mM, 20-30 mM, 15-20 mM, 20-25 mM, 25-30 mM, or 18-22 mM of histidine, citrate, or phosphate. In some embodiments, the formulation comprises 10 mM, 15 mM, 16 mM, 17 mM, 18 mM, 19 mM, 20 mM, 21 mM, 22 mM, 23 mM, 24 mM, 25 mM, 26 mM, 27 mM, 28 mM, 29 mM, or 30 mM histidine, citrate, or phosphate, or a range bounded by two of these concentrations. In certain embodiments, the formulation comprises 20 mM histidine, citrate, or phosphate.

[0100] In some embodiments, the formulation includes histidine as a buffer. In some such embodiments, the formulation does not include citrate or phosphate. In some such cases, the formulation buffer consists essentially of histidine. In some embodiments, the formulation includes citrate as a buffer. In some such embodiments, the formulation does not include histidine or phosphate. In some such cases, the formulation buffer consists essentially of citrate. In some embodiments, the formulation includes phosphate as a buffer. In some such embodiments, the formulation does not include citrate or histidine. In some such cases, the formulation buffer consists essentially of phosphate.

[0101] In some embodiments comprising or consisting essentially of histidine as a buffer, the formulation comprises 5-40 mM histidine, e.g., 10-40 mM, 10-30 mM, 15-25 mM, 10-20 mM, 20-25 mM, 25-30 mM, or 18-22 mM histidine. In some embodiments, the formulation comprises 10 mM, 15 mM, 16 mM, 17 mM, 18 mM, 19 mM, 20 mM, 21 mM, 22 mM, 23 mM, 24 mM, 25 mM, 26 mM, 27 mM, 28 mM, 29 mM, or 30 mM histidine, or a range bounded by two of these concentrations. In some embodiments, the formulation comprises 20 mM histidine.

[0102] In some embodiments, the surfactant is a polysorbate, such as polysorbate 20 or polysorbate 80. In some such cases, the formulation comprises 0.001-0.1%, 0.002-0.1%, 0.01-0.1%, 0.005%-0.05%, 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 or 80. In some embodiments, the formulation comprises 0.005%-0.05% polysorbate 20, such as 0.008%-0.012% polysorbate 20. In other embodiments, the formulation contains 0.002-0.1% polysorbate 20. In some embodiments, the formulation contains 0.01% polysorbate 20.

[0103] Sucrose-containing formulations In some embodiments, the formulation comprises or consists essentially of a mixture of anti-FGFR2 antibody, histidine, polysorbate (e.g., polysorbate 20), and sucrose. The concentrations of the antibody, histidine, and polysorbate can be, for example, as provided above. In some embodiments, the sucrose can be found in an amount that provides a formulation that is isotonic with body fluids. In some embodiments, the formulation comprises 200-300 mM, e.g., 200-250 mM, 250-300 mM, 200 mM, 210 mM, 220 mM, 230 mM, 240 mM, 250 mM, 260 mM, 270 mM, 280 mM, 290 mM, or 300 mM sucrose. In some embodiments, the formulation comprises 250-300 mM sucrose. In other embodiments, the formulation comprises 260-280 mM sucrose. In certain embodiments, the formulation comprises 250-270 mM sucrose. In some embodiments, the formulation comprises 270-300 mM sucrose. In other embodiments, the formulation comprises 250 mM, 260 mM, 270 mM, or 280 mM sucrose. In certain embodiments, the formulation comprises 270 mM sucrose. In some embodiments, the formulation comprising sucrose does not comprise arginine.

[0104] In some embodiments, the pH range of the pharmaceutical formulation is 5.0-7.0. In some embodiments, the sucrose-based pharmaceutical formulation has a pH of 5.0-6.5, such as 5.0-6.0, 5.5-6.0, 5.5-6.5, 5.8-6.2, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, or 6.5. In some embodiments, the pH is 5.8-6.2. In some embodiments, the pH is 6.0.

[0105] Preparations containing arginine In some embodiments, the formulation comprises or consists essentially of histidine, arginine, a polysorbate (e.g., polysorbate 20), and an antibody. In some such embodiments, the formulation does not comprise sucrose.

[0106] In some embodiments, arginine is found at 100-200 mM, such as 100-150 mM, 150-200 mM, 130-170 mM, or 140-160 mM. In certain embodiments, arginine is present at a concentration of 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, or 200 mM. In other embodiments, arginine is found at 140 mM, 150 mM, or 160 mM. In some embodiments, arginine is present at 150 mM.

[0107] In some embodiments, the pH range of the pharmaceutical formulation is 5.0-7.0. In some embodiments, the arginine-based pharmaceutical formulation has a pH of 5.0-6.5, such as 5.0-6.0, 5.5-6.5, 5.5-6.0, 5.5-5.9, 5.6-5.8, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, or 6.5. In some embodiments, the pH is 5.5-5.9. In some embodiments, the pH is 5.6-5.8. In some embodiments, the pH is 5.7.

[0108] Specific Formulation Embodiments In other embodiments, the pharmaceutical formulation comprises 10-30 mg / mL of an anti-FGFR2 antibody, 130-170 mM arginine or 250-290 mM sucrose, 0.002-0.1% polysorbate 20, and one or more buffers selected from 10-30 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.0-7.0. In some such cases, the pharmaceutical formulation comprises 10-30 mg / mL of an anti-FGFR2 antibody, 130-170 mM arginine or 250-290 mM sucrose, 0.002-0.1% polysorbate 20, and one or more buffers selected from 10-30 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.0-7.0.

[0109] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL of an anti-FGFR2 antibody, 130-170 mM arginine or 250-290 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 10-30 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.0-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL of an anti-FGFR2 antibody, 130-170 mM arginine or 250-290 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 10-30 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.0-6.5. In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL of an anti-FGFR2 antibody, 130-170 mM arginine or 250-290 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 10-20 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.0-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL of an anti-FGFR2 antibody, 130-170 mM arginine or 250-290 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 10-30 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.0-6.5.

[0110] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL of an anti-FGFR2 antibody, 140-160 mM arginine or 260-280 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 15-25 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL of an anti-FGFR2 antibody, 140-160 mM arginine or 260-280 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 15-25 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL of an anti-FGFR2 antibody, 140-160 mM arginine or 260-280 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 15-25 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL of an anti-FGFR2 antibody, 140-160 mM arginine or 260-280 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 15-25 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.5-6.5.

[0111] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL of an anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 15-25 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL of an anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 15-25 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL of an anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 15-25 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL of an anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.005-0.05% polysorbate 20, and one or more buffers selected from 15-25 mM histidine, citrate, and phosphate, and the formulation has a pH of 5.5-6.5.

[0112] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.005-0.05% polysorbate 20, and 15-25 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.005-0.05% polysorbate 20, and 15-25 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.005-0.05% polysorbate 20, and 15-25 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.005-0.05% polysorbate 20, and 15-25 mM histidine, and the formulation has a pH of 5.5-6.5.

[0113] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.008-0.012% polysorbate 20, and 10-30 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.008-0.012% polysorbate 20, and 10-30 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.008-0.012% polysorbate 20, and 10-30 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.008-0.012% polysorbate 20, and 10-30 mM histidine, and the formulation has a pH of 5.5-6.5.

[0114] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.008-0.012% polysorbate 20, and 15-25 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.008-0.012% polysorbate 20, and 15-25 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.008-0.012% polysorbate 20, and 15-25 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.008-0.012% polysorbate 20, and 15-25 mM histidine, and the formulation has a pH of 5.5-6.5.

[0115] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.5-6.5. In some embodiments, the pharmaceutical formulation consists essentially of 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine or 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.5-6.5.

[0116] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL of an anti-FGFR2 antibody, 150 mM arginine, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.6-5.8 or 5.7. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL of an anti-FGFR2 antibody, 150 mM arginine, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.6-5.8 or 5.7. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL of an anti-FGFR2 antibody, 150 mM arginine, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.6-5.8 or 5.7. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL anti-FGFR2 antibody, 150 mM arginine, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.6-5.8 or 5.7.

[0117] In some embodiments, the pharmaceutical formulation comprises 10-30 mg / mL of an anti-FGFR2 antibody, 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.9-6.1 or 6.0. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL of an anti-FGFR2 antibody, 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.9-6.1 or 6.0. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL of an anti-FGFR2 antibody, 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.9-6.1 or 6.0. In some embodiments, the pharmaceutical formulation comprises 10-20 mg / mL anti-FGFR2 antibody, 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.9-6.1 or 6.0.

[0118] In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL anti-FGFR2 antibody, 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.0-6.0. In some embodiments, the pharmaceutical formulation consists essentially of 10-30 mg / mL anti-FGFR2 antibody, 270 mM sucrose, 0.01% polysorbate 20, and 20 mM histidine, and the formulation has a pH of 5.5-6.0.

[0119] In some embodiments, the formulation does not include certain types of excipients. For example, in some embodiments, the formulation does not include one or more of sugar alcohols, proteins other than anti-FGFR2 antibodies, surfactants other than polysorbates, and amino acids other than arginine and / or histidine. In certain embodiments that include arginine, the formulation does not include one or more of sugars, sugar alcohols, proteins other than anti-FGFR2 antibodies, surfactants other than polysorbates, and amino acids other than arginine and histidine. In some embodiments that include sucrose, the formulation does not include one or more of sugars other than sucrose, sugar alcohols, proteins other than anti-FGFR2 antibodies, surfactants other than polysorbates, and amino acids other than histidine. In some embodiments that include arginine, the formulation does not include any of sugars, sugar alcohols, proteins other than anti-FGFR2 antibodies, surfactants other than polysorbates, and amino acids other than arginine and histidine. In some embodiments that include sucrose, the formulation does not include any of the following: sugars other than sucrose, sugar alcohols, proteins other than anti-FGFR2 antibodies, surfactants other than polysorbates, and amino acids other than histidine. In certain embodiments, the formulation does not include any other buffer components other than histidine, citrate, and / or phosphate. In certain embodiments that include histidine, citrate, or phosphate as a buffer, the formulation does not include any other buffer components. As stated earlier in this application, the term "free" in this context means that the excluded component is not present beyond trace levels, for example, due to contamination or impurities found in other intentionally added components.

[0120] Exemplary Properties of the Formulation In some embodiments, the formulation is stored as a liquid and is not lyophilized before administration to a patient. In some embodiments, the formulation is a ready-to-use liquid formulation and therefore can be administered directly to a patient. Liquid formulations offer the advantage that they are ready-to-use and easier to administer to a patient than lyophilized formulations. In other embodiments, the formulation is diluted in saline, water, or mixed with other substances to dilute the protein before administration. For example, a formulation designed for IV infusion can be diluted in saline or another suitable buffer in an IV bag before administration. In some embodiments, a liquid formulation, such as a ready-to-use liquid formulation, is administered directly intravenously without dilution.

[0121] In some embodiments, the formulation is administered intravenously, for example by intravenous (IV) infusion. Administration can occur in a hospital, clinic, outpatient clinic, or other medical office setting. IV administration allows for the administration of protein therapeutics in relatively large volumes of liquid.

[0122] In other embodiments, for higher protein (antibody) concentrations (e.g., greater than 300 mg / mL), the pharmaceutical formulations herein are injectable or administered subcutaneously, e.g., by injection, e.g., using a subcutaneous administration device. In some embodiments, the formulation is contained in a vial. In other embodiments, the vial is part of or attached to a subcutaneous administration device, such as a syringe and needle. In some embodiments, the vial is a single-use vial. The vials herein may hold, for example, 0.5 mL, 1 mL, 1.5 mL, 2 mL, or 3 mL of the formulation. In some embodiments, a complete single dose of anti-FGFR2 antibody may be administered from one, two, or three subcutaneous administrations. Thus, in some such embodiments, a complete single dose of the antibody may be contained within one, two, or three single-use vials. In certain embodiments, the vial contains 1-2 mL of the formulation. Thus, in some embodiments, a complete single dose for a patient may be contained in a single 1 mL, 1.5 mL, 2 mL, 2.5 mL, or 3 mL vial, or in two or three 0.5 mL, 1 mL, 0.5 mL, or 2 mL vials. In some embodiments, the liquid formulation is stored in the dark, e.g., in a vial in the dark.

[0123] In some embodiments, the pharmaceutical formulations herein remain stable in solution, e.g., suitable for pharmaceutical use after 3 or 6 months at 40° C. In other embodiments, the pharmaceutical formulations herein remain stable in solution, e.g., suitable for pharmaceutical use after 6 months at 5° C. and / or 25° C. In certain embodiments, protein aggregation in the formulation may not increase by more than 3%, 4%, 5%, 6%, or 7% after 1 month, 2 months, or 3 months of storage at 40° C. In yet other embodiments, protein aggregation in the formulation may not increase by more than 3% or 4% after 1 month of storage at 40° C. and / or may not increase by more than 7% after 3 months of storage at 40° C. In some embodiments, protein aggregation in the formulation may not increase by more than 2% or 2.5% after 3 months of storage at 25° C. In some embodiments, protein aggregation in the formulation may not increase by more than 2% after 6 months of storage at 5° C.

[0124] In addition to aggregates, charged variants of proteins may form during storage as a result of chemical changes such as deamidation and oxidation. For example, in some embodiments, the acidic variants of anti-FGFR2 antibodies do not increase or decrease by more than 20% after 3 months of storage at 25° C. In some embodiments, there is no change in the percentage of charged variants after 6 months of storage at 5° C. The appearance of charged variants can be detected, for example, using cation exchange chromatography or anion exchange chromatography.

[0125] In certain embodiments, the pharmaceutical composition comprises 40% or less of an acidic variant of the anti-FGFR2 antibody and 20% or less of a basic variant of the anti-FGFR2 antibody after storage for 1 month at 40° C. In some embodiments, the pharmaceutical composition comprises 1%-40%, 15%-40%, or 20%-40%, 20%-30%, or 30%-40% of an acidic variant of the antibody and / or 5%-30%, 5%-20%, 10%-20%, 20%-30%, or 15%-20% of a basic variant of the antibody after storage for 1 month at 40° C. In some embodiments, the pharmaceutical composition comprises an anti-FGFR2 antibody, the composition comprises 15%-35%, 20%-30%, or 25%-30% of an acidic variant of the antibody and / or 15%-40%, 15%-30%, 15%-25%, 15%-20%, or 20%-25% of a basic variant of the antibody or antigen-binding fragment thereof after 3 months at 25° C. In some embodiments, the pharmaceutical composition comprises an anti-FGFR2 antibody, the composition comprises 15%-30%, 15%-25%, 20%-30%, or 20%-25% of an acidic variant of the antibody and / or 20%-35%, 20%-30%, or 25%-30% of a basic variant of the antibody or antigen-binding fragment thereof after 6 months at 5° C.

[0126] For example, in some embodiments, a formulation comprising histidine, sucrose, and polysorbate 20 having a pH of 5.5 to 6.5 herein initially contains 20 to 25% acidic variants of an anti-FGFR2 antibody, but after one month of storage at 40° C., it contains 20% to 40% acidic variants, after three months of storage at 25° C., it contains 25% to 30% acidic variants, and after six months of storage at 5° C., it contains 20% to 25% acidic variants (see Figures 27A to C). In some embodiments, a formulation comprising histidine, sucrose, and polysorbate 20 having a pH of 5.5 to 6.5 herein initially contains 20 to 25% acidic variant of an anti-FGFR2 antibody, but after storage at 40° C. for one month, it contains 20% to 40% (e.g., 30% to 40%) acidic variant, after storage at 25° C. for three months, it contains 25% to 30% acidic variant, and after storage at 5° C. for six months, it contains 20% to 25% acidic variant (see FIGS. 28A-C). In some embodiments, a formulation comprising histidine, sucrose, polysorbate 20 at pH 5.5-6.5 herein initially contains 30% basic variant of an anti-FGFR2 antibody, but after one month of storage at 40° C., it contains 10%-25% basic variant, after three months of storage at 25° C., it contains 15%-30% (e.g., 15%-25%) basic variant, and after six months of storage at 5° C., it contains 25%-30% basic variant (see Figures 29A-C). In some embodiments, a formulation comprising histidine, arginine, polysorbate 20 and having a pH of 5.5-6.5 herein initially contains 30-35% basic variants of an anti-FGFR2 antibody, but after one month of storage at 40° C., contains 10%-20% basic variants, after three months of storage at 25° C., contains 15%-30% (e.g., 15%-25%) basic variants, and after six months of storage at 5° C., contains 25%-30% basic variants (see Figures 30A-C).

[0127] In some embodiments, the pharmaceutical formulation remains stable for pharmaceutical use after one or more freeze-thaw cycles, for example, 2, 3, 4, or 5 freeze-thaw cycles, for example, at -70°C. For example, in some embodiments, protein aggregation in the formulation may not increase by more than 2.0%, 1.5%, 1.0%, or 0.5% after 5 freeze-thaw cycles. In certain embodiments, the percentage of high molecular weight protein species measured by light scattering remains unchanged or does not increase by more than 0.5% after 5 freeze-thaw cycles. In some embodiments, the formulation also remains stable to mechanical stress, which may be determined by exposing a vial containing the formulation to agitation for an extended period of time. In other embodiments, aggregation may not increase by more than 2.0%, 1.5%, 1.0%, or 0.5% after 72 hours of mechanical stress at 500 rpm.

[0128] Therapeutic methods using anti-FGFR2 antibody pharmaceutical preparations The pharmaceutical formulations disclosed herein can be used in methods for treating patients in need of anti-FGFR2 antibody therapy. Use of anti-FGFR2 antibodies is disclosed, for example, in International Patent Publication Nos. WO2015 / 017600 and WO2017 / 091577, each of which is incorporated herein by reference.

[0129] In some embodiments, the pharmaceutical formulations herein can be used to treat cancer patients. In some embodiments, the cancer is a solid tumor. More specific non-limiting examples of such cancers include squamous cell carcinoma, small cell lung cancer, pituitary cancer, esophageal cancer, astrocytoma, soft tissue sarcoma, non-small cell lung cancer (including squamous cell non-small cell lung cancer), adenocarcinoma of the lung, squamous cell carcinoma of the lung, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatocellular carcinoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, renal cancer, renal cell carcinoma, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, liver cancer, brain cancer, endometrial cancer, testicular cancer, bile duct cancer, gallbladder cancer, gastric cancer, melanoma, and various types of head and neck cancer (including squamous cell carcinoma of head and neck cancer). In some embodiments, the cancer is gastric cancer or bladder cancer.

[0130] In some embodiments, the cancer is recurrent or progressive after a therapy selected from surgery, chemotherapy, radiation therapy, or a combination thereof. In some embodiments, the subject is a poor responder to a PD-1 / PD-L1 inhibitor. A subject who is a poor responder to a PD-1 / PD-L1 inhibitor is a subject who has previously responded to a PD-1 / PD-L1 inhibitor but may have become less responsive to a PD-1 / PD-L1 inhibitor, or a subject who may have never responded to a PD-1 / PD-L1 inhibitor. In some embodiments, the subject has previously received a PD-1 / PD-L1 inhibitor therapy.

[0131] In some embodiments, the cancer comprises FGFR2 gene amplification. In some embodiments, the cancer comprising FGFR2 gene amplification also overexpresses FGFR2IIIb. In some embodiments, the cancer comprising FGFR2 amplification overexpresses FGFR2IIIb to a greater extent than FGFR2IIIc. In some embodiments, the cancer comprising FGFR2 amplification expresses FGFR2IIIb at a normalized level that is 2-fold, 3-fold, 5-fold, or more than 10-fold greater than the normalized level of FGFR2IIIc expression. In some embodiments, the expression level is normalized to GUSB. In some embodiments, the cancer overexpresses FGFR2IIIb but does not comprise FGFR2 gene amplification.

[0132] In some embodiments, the cancer is a gastric cancer comprising FGFR2 gene amplification. In some embodiments, the gastric cancer comprising FGFR2 gene amplification overexpresses FGFR2IIIb. In some embodiments, the gastric cancer comprising FGFR2 amplification overexpresses FGFR2IIIb to a greater extent than FGFR2IIIc. In some embodiments, the gastric cancer comprising FGFR2 amplification expresses FGFR2IIIb at a normalized level that is 2-fold, 3-fold, 5-fold, or more than 10-fold greater than the normalized level of FGFR2IIIc expression. In some embodiments, the expression level is normalized to GUSB. In some embodiments, the gastric cancer overexpresses FGFR2IIIb but does not comprise FGFR2 gene amplification. In some embodiments, the overexpression is mRNA overexpression. In some embodiments, the overexpression is protein overexpression.

[0133] FGFR2IIIb gene amplification can be determined by any suitable method in the art, including but not limited to in situ hybridization (ISH).In some embodiments, FGFR2 amplification comprises a FGFR2:CEN10 (chromosome 10 centromere) ratio of greater than 3.

[0134] FGFR2IIIb mRNA overexpression can be determined by any suitable method in the art, including but not limited to quantitative PCR (qPCR). The term "FGFR2IIIb mRNA overexpression" refers to elevated levels of FGFR2IIIb mRNA, regardless of the cause of such elevated levels (i.e., whether the elevated levels are the result of increased transcription and / or decreased degradation of mRNA, other mechanisms, or a combination of mechanisms).

[0135] FGFR2IIIb protein overexpression can be determined by any suitable method in the art, including, but not limited to, antibody-based methods such as immunohistochemistry (IHC). In some embodiments, IHC staining is scored according to methods in the art. The term "FGFR2IIIb protein overexpression" refers to an increase in the level of FGFR2IIIb protein, regardless of the cause of such increase in level (i.e., whether the increase in level is the result of increased translation and / or decreased degradation of the protein, other mechanisms, or a combination of mechanisms). In some embodiments, 1+, 2+, or 3+ staining of tumor cells by IHC indicates FGFR2IIIb overexpression. For example, overexpression can be determined by a 1+, 2+, or 3+ IHC signal in at least 10% of tumor cells, e.g., at least 20%, 30%, 40%, or 50% of tumor cells. In some embodiments, 2+ or 3+ staining of tumor cells by IHC indicates FGFR2IIIb overexpression. For example, overexpression can be determined by an IHC signal of 2+ or 3+ in at least 10% of the tumor cells, such as at least 20%, 30%, 40%, or 50% of the tumor cells.

[0136] In some embodiments, FGFR2 overexpression may be reported as an "H score." For example, in some such embodiments, the tumor is a bladder cancer tumor. To determine the H score, a first membrane staining intensity may be determined for cells in a fixed field, e.g., via IHC, to obtain a score of 0, 1+, 2+, or 3+, and the H score may be calculated using the following formula: 1×(% of cells visualized with an IHC intensity of 1+)+2×(% of cells visualized with an IHC intensity of 2+)+3×(% of cells visualized with an IHC intensity of 3+). Theoretically, the H score may range from 0 to 300, and is equal to 300 if all cells in the field have an IHC staining of 3+. In some embodiments, the patient to be treated has a starting H-score for FGFR2, such as FGFR2b (e.g., FGFR2IIIb) of >20, e.g., >30, >40, >50, or >100, or in the range of 20-300, 20-100, 20-50, 20-40, or 20-30. In some embodiments, the patient has an H-score of >10, or in the range of 10-20 or 15-20. In other embodiments, the patient has an H-score of 0-10, which may indicate a lack of FGFR2 overexpression. In some such embodiments, the patient is a bladder cancer patient.

[0137] In some embodiments where the patient is afflicted with gastric or bladder cancer, the subject may have previously been determined to have one of the following profiles, or the treatment method may include determining whether the patient fits one of the following profiles with respect to FGFR2 expression / gene amplification, which may indicate a level of predicted responsiveness to treatment: a) for gastric cancer subjects, an IHC signal of 3+ in at least 10% of the tumor cells, b) for gastric cancer subjects, an IHC signal of 3+ in at least 10% of the tumor cells, and FGF Amplification of the R2 gene, c) for gastric cancer subjects, an IHC signal of 3+ in at least 10% of the tumor cells without amplification of the FGFR2 gene, d) for gastric cancer subjects, an IHC signal of 1+ or 2+ in at least 10% of the tumor cells, e) for bladder cancer subjects, an IHC signal of 1+ in at least 10% of the tumor cells, f) for bladder cancer subjects, an IHC signal of 2+ in at least 10% of the tumor cells, g) for bladder cancer subjects, an H score of greater than 20, h) for bladder cancer subjects, an H score of 10-19, i) for bladder cancer subjects, an H score of less than 10. In some embodiments where the patient is afflicted with gastric or bladder cancer, the subject may have previously been determined to have one of the following profiles, or the treatment method may include determining whether the patient fits one of the following profiles with respect to FGFR2 expression / gene amplification, which may indicate a level of predicted responsiveness to treatment: a) for gastric cancer subjects, an IHC signal of 3+ in at least 5% of the tumor cells; b) for gastric cancer subjects, an IHC signal of 3+ in at least 5% of the tumor cells; and c) for gastric cancer subjects, an IHC signal of 3+ in at least 5% of the tumor cells; amplification of the FGFR2 gene, c) for gastric cancer subjects, an IHC signal of 3+ in at least 5% of tumor cells without amplification of the FGFR2 gene, d) for gastric cancer subjects, an IHC signal of 1+ or 2+ in at least 5% of tumor cells, e) for bladder cancer subjects, an IHC signal of 1+ in at least 5% of tumor cells, f) for bladder cancer subjects, an IHC signal of 2+ in at least 5% of tumor cells, g) for bladder cancer subjects, an H score of greater than 20, h) for bladder cancer subjects, an H score of 10-19, i) for bladder cancer subjects, an H score of less than 10.In some embodiments where the patient is suffering from gastric or bladder cancer, the subject may have previously been determined to have one of the following profiles, or the treatment method includes determining whether the patient fits one of the following profiles with respect to FGFR2 expression / gene amplification, which may indicate a level of expected responsiveness to treatment: a) the cancer is gastric cancer with a 2+ or 3+ FGFR2-IIIb immunohistochemistry (IHC) signal in the cancer sample, b) the cancer is gastric cancer with a 2+ or 3+ FGFR2-IIIb IHC signal in the cancer sample, and the FGFR2 gene is amplified, c) the cancer is gastric cancer with a 2+ or 3+ FGFR2-IIIb IHC signal in the cancer sample, and the FGFR2 gene is not amplified, or d) the cancer is bladder cancer with a 2+ or 3+ FGFR2-IIIb IHC signal in the cancer sample.

[0138] In some embodiments, the anti-FGFR2 antibody may be administered to a cancer patient at a dose of at least 0.1, 0.3, 0.5, 1, 2, 3, 4, 5, 10, 15, 20, 25, or 30 mg / kg, or within a range bounded by any two of these doses. In some embodiments, the antibody formulation is administered once every 1, 2, 3, 4, or 5 weeks.

[0139] In some embodiments, pharmaceutical formulations comprising anti-FGFR2 antibodies are provided as part of a combination of drugs, such as immune checkpoint inhibitors (immunostimulants) and chemotherapeutic agents. For example, anti-FGFR2 antibodies or formulations comprising them can be provided before, substantially simultaneously with, or after other treatment modalities, such as surgery, chemotherapy, or radiation therapy. In some embodiments, an effective amount of the formulations herein comprising anti-FGFR2 antibodies is administered in conjunction with another anti-cancer agent. Non-limiting exemplary anti-cancer drugs that may be administered with the anti-FGFR2 antibody include platinum agents (such as cisplatin, oxaliplatin, and carboplatin), paclitaxel (TAXOL®), albumin engineered nanoparticle formulation of paclitaxel (ABRAXANE®), docetaxel (TAXOTERE®), gemcitabine (GEMZAR®), capecitabine (XELODA®), irinotecan (CAMPTOSAR®, epirubicin (ELLENCE®, PHARMORUBICIN®), FOLFOX (oxaliplatin in combination with 5-FU and leucovorin), FOLFIRI (leucovorin, 5-FU and irinotecan), and ribavirin (RIBA). In some embodiments, an effective amount of an anti-FGFR2 antibody formulation is administered in combination with paclitaxel. In some embodiments, an effective amount of an anti-FGFR2 antibody formulation is administered in combination with cisplatin and / or 5-FU. In some embodiments, an effective amount of an anti-FGFR2 antibody formulation is administered in combination with FOLFOX (oxaliplatin, 5-FU, and leucovorin).

[0140] In some embodiments, a method for treating cancer is provided that includes administering an effective amount of a pharmaceutical formulation comprising an anti-FGFR2 antibody and at least one immunostimulant. In some other embodiments, a method for treating cancer is provided that includes administering a formulation comprising an effective amount of an anti-FGFR2 antibody and an effective amount of at least one immunostimulant. In an exemplary embodiment, the at least one immunostimulant comprises a PD-1 / PD-L1 inhibitor. In some embodiments, the formulation comprising anti-FGFR2 and at least one immunostimulant, such as a PD-1 / PD-L1 inhibitor, are administered simultaneously. In some embodiments, the formulation comprising anti-FGFR2 antibody and at least one immunostimulant, such as a PD-1 / PD-L1 inhibitor, are administered sequentially. In some embodiments, at least 1, at least 2, at least 3 doses, at least 5 doses, or at least 10 doses of an anti-FGFR2 antibody are administered prior to administration of at least one immunostimulant, such as a PD-1 / PD-L1 inhibitor. In some embodiments, at least 1, at least 2, at least 3 doses, at least 5 doses, or at least 10 doses of at least one immune stimulant, such as a PD-1 / PD-L1 inhibitor, are administered prior to administration of the formulation comprising an anti-FGFR2 antibody. In some embodiments, the final dose of at least one immune stimulant, such as a PD-1 / PD-L1 inhibitor, is administered at least 1 day, 2 days, 3 days, 5 days, or 10 days, or 1 week, 2 weeks, 3 weeks, 5 weeks, 12 weeks, or 24 weeks, prior to the first dose of the anti-FGFR2 antibody. In some other embodiments, the final dose of the anti-FGFR2 antibody is administered at least 1 day, 2 days, 3 days, 5 days, or 10 days, or 1 week, 2 weeks, 3 weeks, 5 weeks, 12 weeks, or 24 weeks prior to the first dose of the at least one immune stimulant, such as a PD-1 / PD-L1 inhibitor. In some embodiments, the subject has undergone or is undergoing PD-1 / PD-L1 inhibitor therapy, and the anti-FGFR2 antibody is added to the treatment regimen. EXAMPLES

[0141] The examples discussed below are intended to be purely illustrative of the present invention and should not be construed as limiting the present invention in any manner. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperatures, etc.), but some experimental error and deviation should be accounted for.

[0142] Example 1: Materials and methods used in formulation studies I. Antibody Production The anti-FGFR2-IIIb antibody used in the following examples was produced in a Chinese Hamster Ovary (CHO) cell line lacking the FUT8 gene (α1,6-fucosyltransferase), and various lots that underwent alterations in the fermentation and purification process were used in the development of the antibody. The anti-FGFR2-IIIb antibody is an afucosylated humanized IgG1 monoclonal antibody. A knowledge-based formulation development approach was used to identify the appropriate composition that provides maximum stability to the protein. This takes into account both the intrinsic properties of the molecule and exogenous formulation components that may affect protein stability. Testing was performed to identify the appropriate components to produce a 10-20 mg / mL liquid formulation that can be diluted for IV infusion.

[0143] Raw Materials: L-Histidine (PN H3911), L-Histidine.HCl (PN H5659) and Sodium Chloride (PN S9888) were obtained from Sigma Aldrich. L-Arginine.HCl (PN 2067-06), Citric Acid (PN 0119), Sodium Citrate (PN 3627) and Polysorbate 20 (PN4116) were obtained from JT Baker (Thermo Fischer Scientific).

[0144] Containers / Closures: The formulations were filled using 3 cc Type I West Pharmaceutical glass vials (PN 68000368) and 13 mm stoppers (PN 19700116), polypropylene tubing, and glass HPLC vials from West Pharmaceutical Services, Inc. (PA).

[0145] II. General Formulation Procedure The formulations were prepared by dialysis of the drug substance into the target formulation buffer using a MWCO 20 kD dialysis membrane. The formulations were filtered in a laminated flow hood using a 0.2 μm filter unit and filled into appropriate container / closure systems for stability evaluation. Individual vials were removed from the designated storage conditions at pre-determined time points for analysis.

[0146] III.Analysis method Stability samples were analyzed using the early development assays described below.

[0147] Visual Inspection: Visual evaluation was performed under fluorescent lighting against a black / white background.

[0148] Protein concentration: Protein concentration was determined by UV absorbance at 280 nm using a theoretical absorption coefficient of 1.43 cm-1 [g / L]-1. Samples were diluted into the linear absorbance range with the appropriate formulation buffer and measured against Dulbecco's Phosphate Buffered Saline (DPBS) water. OD measurements were performed on a Beckman Coulter DU800 UV-Vis spectrophotometer (Beckman Coulter, Inc., CA).

[0149] pH: Buffer pH was determined using a calibrated Beckman Coulter pHi560 pH meter (Beckman Coulter, Inc., CA).

[0150] Osmolality: Buffer osmolality was measured by vapor pressure using a Wescor VAPRO® system (Wescor, Inc., UT).

[0151] Differential Scanning Calorimetry (DSC) Analysis: High-sensitivity Differential Scanning Calorimetry (HSDSC) analysis was performed using a VP-DSC (MicroCal, Northampton, MA). Approximately 0.5 mg of each sample was loaded onto a sample cell and an equal volume of matching dialysis buffer was loaded onto a reference cell. Samples were scanned from 20 to 90°C or 100°C at a rate of 1°C / min. Data were analyzed with Origin 7.0 data analysis software (OriginLab, MA).

[0152] Ion exchange chromatography (IEX): Cation exchange chromatography is a method to separate protein variants based on their net surface charge. The isoelectric point of a protein (the pH at which a protein has no net charge, pI) is determined by its primary amino acid sequence, while the net charge of a protein variant is based on the protein's isoelectric charge and the running buffer pH. For example, at a running buffer pH of 7, approximately one unit lower than the pI of an antibody of 8.4, the protein carries a net positive charge and electrostatically cross-links to the oppositely weakly charged acidic carboxylate functional groups located on a Propac WCX-10 column resin with a substrate of ethylvinylbenzene-divinylbenzene. Using a high performance liquid chromatography system with UV detection at 280 nm, the protein variants were separated and eluted from the column when applying an increasing salt gradient, with the variants with the weakest ionic interactions starting to elute first and the variants with stronger ionic interactions eluting later at higher salt concentrations. The isoforms were sequentially eluted into three regions as acidic, main, and basic, with the highest peak being the main isoform peak. Based on the peak area, each area had a calculated relative peak percentage. A weak cation exchange HPLC method was performed using a Dionex WCX-10 ProPac™ 4X250 mm column (Thermo Fischer Scientific). Samples were run on an Agilent 1200 series HPLC and chromatograms were integrated using Chemstation software (Agilent Technologies, CA). Mobile phase A was 10 mM HEPES at pH 7.0 and mobile phase B was 10 mM HEPES, 100 mM sodium chloride at pH 7.0. A gradient method from 40-90% B in 28 min was used with UV detection at 280 nm. The percentages of acidic, basic, and main peaks were reported from the total area percent of each peak-associated absorbance.

[0153] Size Exclusion Chromatography (SEC): Size exclusion chromatography is a method to separate proteins based on their size. The method relies on a porous resin to separate molecules of different sizes. Large molecules elute early because these molecules cannot access the pores of the resin and pass through the column unhindered. Smaller molecules are able to access the resin pores and increase their path length through the column due to distortions within the resin particles. Thus, smaller molecules elute after larger molecules. Two methods were used for size exclusion chromatography. The first method utilized a Tosoh G3000SWXL 7.8 x 300 mm column (Sigma Aldrich, Inc., MO) with 100 mM sodium phosphate, 700 mM arginine (pH 6.8) as the mobile phase at 0.5 mL / min for 30 minutes. The second method utilized a Sepax Zenix SEC-300 7.8 x 200 mm column (Sepax Technologies, Inc., Delaware) using 100 mM sodium phosphate, 400 mM sodium chloride (pH 6.8) as the mobile phase at 1 mL / min for 12 min. Samples were run on an Agilent 1100 & 1200 series HPLC and chromatograms were integrated using Chemstation software with detection set at 280 nm. Percentages of aggregates, low molecular weight (LMW), and main peaks were reported from the total area percent of absorbance associated with each peak.

[0154] Light scattering: Using a spectrophotometer, 280 nm (UV A 280 ) was measured. The concentration of the antibody samples was calculated using Beer's law, A = εlc (ε = extinction coefficient, l = length of solution through which light passes (path length of the cuvette) (cm), and c = concentration of the solution). The theoretical extinction coefficient was used to determine the antibody concentration (e.g., the theoretical extinction coefficient for anti-FGFR2 antibody is 1.43 (OD mL) / (mg cm)). The concentration (mg / mL) of each sample preparation was calculated as follows: Concentration = (Absorbance at A280 - Absorbance at A350) x (Dilution factor) / (Extinction coefficient x Path length). The mean and standard deviation of each sample were calculated.

[0155] Example 2: Water Soluble The anti-FGFR2-IIIb antibody was concentrated to approximately 100 mg / mL and dialyzed against water. The sample was then further concentrated to 180 mg / mL. The anti-FGFR2-IIIb antibody was water soluble at 180 mg / mL. Maximum solubility was not reached.

[0156] Example 3: Initial freeze / thaw stability of anti-FGFR2-IIIb antibodies Freeze / thaw studies were performed on anti-FGFR2-IIIb antibody. The antibody was concentrated to 11.6 mg / mL and formulated in 1×PBS at pH 7.4 with or without 0.01% polysorbate 20. The formulated solution was filled into vials and subjected to up to three cycles of freeze / thaw from -70°C to ambient temperature. Sample vials were analyzed by visual inspection and SE-HPLC. The freeze / thaw results are shown in Table 1. A slight increase in aggregates was observed over three freeze / thaw cycles. The results suggest that 1×PBS is acceptable for the formulation. TIFF2024138284000002.tif144168

[0157] Example 4: Oxidation test of anti-FGFR2-IIIb antibody The anti-FGFR2-IIIb antibody has a total of 10 methionine residues, of which 4 methionines are in each of the heavy chains, none of which are located in CDR1, and in addition, one methionine is present in the light chain.

[0158] Experiments were performed to evaluate the susceptibility of anti-FGFR2-IIIb antibodies to methionine oxidation by incubating them with 0.01%, 0.1%, or 1.0% hydrogen peroxide overnight at room temperature. The oxidized samples were then characterized by trypsin mapping LC / MS to qualitatively determine the oxidation level of each methionine residue. The activity of the oxidized samples was determined by a cell-based assay for potency, and the binding affinity was analyzed by Biacore™ (GE Healthcare). The peptide map LC / MS characterization shown in Table 2 revealed that light chain methionine 5 and heavy chain methionine 81 were less susceptible to oxidation. Heavy chain methionines 249, 355, and 425 were more susceptible to oxidation. All of the oxidized samples showed comparable potency to the control by ELISA assay. TIFF2024138284000003.tif95170

[0159] Binding of anti-FGFR2-IIIb antibody to FGFR2b and protein A was measured by Biacore. Oxidation of heavy chain methionine did not prevent binding of anti-FGFR2-IIIb antibody to FGFR2b (Figure 1 and Table 3). Increased oxidation prevented binding to protein A. Oxidative degradation of anti-FGFR2-IIIb antibody was not significant considering that the activity of anti-FGFR2-IIIb antibody was not significantly affected in highly oxidized samples. TIFF2024138284000004.tif115170

[0160] Example 5: Effect of buffer pH on protein stability Formulation pH influences protein stability. Formulation pH can affect biochemical degradation pathways such as deamidation, isomerization, and oxidation, as well as biophysical degradation such as aggregation and fragmentation due to interactions of proteins with their environment. Buffer pH was studied as a formulation variable that can affect product stability. The effect of pH on anti-FGFR2-IIIb antibody conformational and chemical stability upon storage was evaluated. Anti-FGFR2-IIIb antibody was purified in two steps and formulated into nine different isotonic formulations (formulations 1-9 in Table 4) with pH values ​​ranging from 4.0 to 8.0. The protein concentration of the buffer exchanged solution was adjusted to 1 mg / mL, and the solution was filled into vials for DSC analysis and isothermal stability evaluation at 40 °C and 25 °C. The formulations evaluated are listed in Table 4. TIFF2024138284000005.tif127163

[0161] Conformational stability by differential scanning calorimetry (DSC) analysis DSC thermograms of formulations 1-9 from Table 4 were generated and analyzed. Formulations 1-3 are marked with * in Table 5 and the results are shown in Figure 2. Three transitions were observed when the samples were heated. After the unfolding transitions, the IgG molecules formed insoluble aggregates and precipitated from the solution. Table 5 shows the unfolding temperature (T M 1) increases with increasing pH, reaching a maximum T of approximately 69°C at pH 6 and pH 7. M The results show that the pH reached 1. The type of buffer did not show a significant effect on the development temperature. TIFF2024138284000006.tif135169

[0162] Isothermal stability The effect of buffer pH on the formation of aggregates, clips (fragments), and charged variants was assessed by monitoring sample stability under accelerated storage conditions at 40°C and 25°C.

[0163] The changes in aggregates and clips were determined by SE-HPLC. Representative SE-HPLC chromatograms of pH screening samples of selected anti-FGFR2-IIIb antibodies are shown in Figure 3. The main peak represents the monomer, the peaks eluting earlier than the monomer are aggregates, and the peaks eluting later are clips. Soluble aggregates increased rapidly in the pH 4 formulation at 40°C, and a moderate increase in soluble aggregates was seen in pH 5.0 and pH 8.0 buffers. A similar effect of pH on clip formation was observed.

[0164] The effect of buffer pH on aggregates (Figure 4) and clips (Figure 5) at 40°C was analyzed by SE-HPLC. Formulations 1, 2, 3, 7, 8, and 9 listed in Table 5 were tested. Within the pH range tested, clip formation was most significant at pH 4. At neutral pH, as shown in Figure 5, clip formation was negligible even after 1 month of storage at 40°C. No obvious increase in aggregates was observed in formulations at pH 5 to pH 8 after at least 2 months of storage at 25°C (Figure 6). No obvious increase in clips was observed in formulations at pH 5 to pH 8 after at least 2 months of storage at 25°C (Figure 7).

[0165] Changes in charge variant distribution are another common degradation pathway for antibodies and are typically related to formulation buffer pH. Charge variants of anti-FGFR2-IIIb antibodies were analyzed by weak cation exchange HPLC (WCX-HPLC) method. A representative chromatogram is shown in Figure 8. The peaks appearing before the main peak are acidic species, and the peaks appearing after the main peak are basic species. The increase in acidic species is due to deamidation of asparagine in the IgG molecule. The change in the charge variant profile occurred dramatically at 40°C. Therefore, the change in the charge profile was monitored at a lower temperature such as 25°C. Figures 9-11 show the effect of buffer pH on the charge variant at 25°C as determined by weak cation exchange high performance liquid chromatography (WCX-HPLC). As shown in Figure 9, there was a rapid increase in acidic species in basic pH conditions, especially at pH 8.0. Samples formulated between pH 5 and pH 6 remained stable even after 2 months of storage at 25°C. Figure 10 shows the effect of pH on the percentage of basic variants for 0-2.5 months of storage at 25° C. Figure 11 shows the effect of pH on the main peak for 0-2.5 months of storage at 25° C.

[0166] The results of the pH screening study indicate that under stress storage conditions, the stability of the anti-FGFR2-IIIb antibody was strongly dependent on the formulation pH. At basic pH, aggregation and formation of acidic variants were the major degradation pathways. The anti-FGFR2-IIIb antibody was determined to be most stable in the pH range of 5.0 to 6.0.

[0167] Example 6: Effect of excipients on protein stability Formulation excipients, such as buffer types and bulking agents, were analyzed for their effect on product stability. To evaluate the effect of excipients on the stability of anti-FGFR2-IIIb antibody, various citrate, phosphate, and histidine buffers were tested. The seven different isotonic solutions buffered at pH 6.0 that were tested are listed in Table 6. TIFF2024138284000007.tif107166

[0168] The protein concentration was adjusted to 20 mg / mL and the solution was filled into vials. Formulations 1-4 in Table 6 were analyzed by DSC. Physical stability was evaluated for all formulations and isothermal stability evaluations at 40° C., 25° C., and 5° C. were determined for formulations 1-5.

[0169] Conformational stability by DSC analysis The unfolding temperatures of various formulations of anti-FGFR2-IIIb antibody were determined by DSC analysis and are summarized in Table 7. After heating to 90°C, precipitation was observed in all formulations. The surfactants in the formulations did not protect the antibody from precipitation after denaturation. As shown in Table 7, the formulations had comparable melting temperatures. TIFF2024138284000008.tif105169

[0170] Three temperature transitions were detected when various formulations of anti-FGFR2-IIIb antibody were heated: anti-FGFR2-IIIb antibody started to unfold at 52.7°C in pH 4 formulation, the unfolding temperature (Tm) increased with increasing pH, reaching a maximum Tm1 at approximately 70°C at pH 6 and pH 7, and the Tm slightly decreased at pH 8.

[0171] physical stability Because protein pharmaceuticals are prone to aggregation when exposed to shear stress, a physical stability study was conducted to evaluate the effect of excipients on the stability of anti-FGFR2-IIIb antibody against multiple freeze / thaw cycles and vigorous agitation. Stability was evaluated by visual observation, A350 light scattering, and SE-HPLC for insoluble and soluble aggregates. The antibody concentration in the samples was 20 mg / mL.

[0172] Agitation stress was applied to the samples by placing the sample vials horizontally on an orbital shaker and shaking the samples at 500 RPM for 77 hours at room temperature. To evaluate the effect of polysorbate 20 on preventing aggregate formation, a direct comparison of arginine formulations with 0%, 0.01%, 0.05%, and 0.10% polysorbate 20 was performed. As shown in Table 8 and Figure 12, no obvious increase in soluble aggregates was detected by SE-HPLC in all formulations.

[0173] Freeze / thaw tests were also performed by freezing samples at -70°C and thawing at ambient temperature for five cycles. As shown in Figure 13, the only obvious change observed in soluble aggregates in the histidine / NaCl formulation was an increase with increasing freeze / thaw cycles. The addition of 0.01% polysorbate 20 to the formulation improved the stability of anti-FGFR2-IIIb antibody against shaking stress. TIFF2024138284000009.tif239170

[0174] Isothermal stability The effect of buffer type and bulking agent on the formation of aggregates, clips, and charged variants was evaluated by monitoring the stability of anti-FGFR2-IIIb antibody under accelerated storage conditions at 40° C., 25° C., and 5° C. The changes in aggregates and clips were monitored by SE-HPLC. The following formulations were tested: 20 mM citrate, 150 mM L-arginine, and 0.01% polysorbate 20; 20 mM phosphate, 150 mM L-arginine, and 0.01% polysorbate 20; 20 mM histidine, 150 mM L-arginine, and 0.01% polysorbate 20; 20 mM histidine, 150 mM NaCl, and 0.01% polysorbate 20; and 20 mM histidine, 150 mM L-arginine, no polysorbate 20.

[0175] As shown in Figure 14, aggregates formed at a higher rate in the citrate and phosphate formulations at 40° C. No significant fragmentation was observed in all formulations after one month of storage at 40° C. All formulations showed the same increase in aggregation after three months of storage at 25° C. (Figure 15).

[0176] The formulation excipients did not affect the charge profile of the anti-FGFR2-IIIb antibody upon storage. As shown in Figure 16, there was an increase in acidic variants in all formulations. Figure 17 shows that there was a decrease in basic variants in all formulations over time, and Figure 18 shows the effect of excipients over time on the main peak.

[0177] Based on antibody stability studies, L-histidine did not significantly affect antibody stability among the buffer types evaluated. Sodium citrate and sodium phosphate had an increase in aggregates after 1 month at 40° C. Thus, L-histidine was more stable than sodium citrate or sodium phosphate, and aggregates increased at a faster rate in the citrate and phosphate formulations.

[0178] Among the bulking agents evaluated, NaCl led to aggregate formation induced by exposure to vigorous shaking and freeze / thaw processes. The most stable antibody profile was obtained with the L-arginine formulation.

[0179] Example 7: pH Evaluation Studies The effect of pH on the chemical and physical stability of the formulations was studied. The following formulations were tested at pH 5.5, 6.0, and 6.5: (a) 20 mM histidine, 150 mM arginine, 0.01% polysorbate 20, and (b) 20 mM histidine, 270 mM sucrose, 0.01% polysorbate 20 (Table 9). The formulations were adjusted to 20 mg / mL and filled into glass vials. TIFF2024138284000010.tif139168

[0180] Effect of pH on physical stability Stability to multiple freeze / thaw cycles and vigorous agitation was evaluated. Stability was assessed by visual observation, A350 light scattering, and SE-HPLC for insoluble and soluble aggregates.

[0181] I. Mechanical Agitation Agitation stress was applied to the samples by placing the sample vials horizontally on an orbital shaker and shaking the samples at 500 RPM for 72 hours at room temperature. No obvious increase in soluble aggregates was detected by SE-HPLC in all formulations. Figure 19 shows the effect of mechanical stress on aggregation of histidine / arginine and histidine / sucrose formulations between pH 5.5-6.5 (pH 5.5, 6.0 and 6.5) over time. Figure 19 shows the effect of mechanical stress on aggregation of histidine / arginine and histidine / sucrose formulations at pH 5.5-6.5. After 72 hours of mechanical agitation, there was no significant change in concentration or light scattering at A350 nm (shown in Table 10). TIFF2024138284000011.tif78168

[0182] II. Freeze-thaw stability Freeze / thaw studies were performed by freezing samples at -70°C for 5 cycles and thawing at ambient temperature. Figures 21 and 22 show the effect of freeze-thaw on aggregation and clipping of histidine / arginine and histidine / sucrose formulations between pH 5.5 and 6.5. In Figure 21, no obvious increase in soluble aggregates was detected by SE-HPLC in either histidine / arginine or histidine / sucrose formulations between pH 5.5 and 6.5. In Figure 22, no obvious increase in clipping was detected by SE-HPLC in either formulation.

[0183] TIFF2024138284000012.tif101168

[0184] The concentration and light scattering remained unchanged after freeze-thaw.

[0185] I. Effect of pH on isothermal stability The effect of buffer pH on the formation of aggregates, clips, and charged variants was assessed by monitoring sample stability under accelerated storage conditions at 40° C., 25° C., and real-time stability at 5° C. The following formulations were tested: (A) 20 mM histidine, 150 mM arginine, 0.01% polysorbate 20 (the "histidine / arginine formulation"), and (B) 20 mM histidine, 150 mM sucrose, 0.01% polysorbate 20 (the "histidine / sucrose formulation").

[0186] In the histidine / arginine and histidine / sucrose formulations, aggregation and clip formation increased with pH after 1 month at 40° C. and after 3 months at 25° C. The histidine / sucrose formulation shows a slightly lower aggregation rate. After 6 months at 5° C., aggregation remained unchanged and clip formation increased by about 1% in both the histidine / arginine and histidine / sucrose formulations.

[0187] For example, Figures 23A-C show aggregate formation in histidine / arginine formulations for various pH conditions measured over (A) 1 month at 40° C., (B) 3 months at 25° C., and (C) 6 months at 5° C. Figures 24A-C show clip formation in histidine / arginine formulations at various pH conditions (pH 5.5-6.5) measured over (A) 1 month at 40° C., (B) 3 months at 25° C., and (C) 6 months at 5° C.

[0188] Figures 25A-C show aggregate formation in histidine / sucrose formulations at various pH conditions measured over (A) 1 month at 40° C., (B) 3 months at 25° C., and (C) 6 months at 5° C. Figures 26A-C show antibody clip formation in histidine / sucrose formulations for various pH conditions (pH 5.5-6.5) measured over (A) 1 month at 40° C., (B) 3 months at 25° C., and (C) 6 months at 5° C.

[0189] Charge isoforms also remained unchanged after 6 months at 5° C. For example, Figures 27A-C show that acidic variants increased in histidine / sucrose formulations after (A) 1 month at 40° C., (B) 3 months at 25° C., and (C) 6 months at 5° C.

[0190] In addition, Figures 28A-C show the effect of pH between pH 5.5-6.5 on acidic variant formation in a histidine / arginine formulation containing 20 mM histidine, 150 mM arginine, and 0.01% polysorbate 20. Figure 28A shows acidic variant formation at pH 5.5-6.0, showing that the % acidic variant remained generally below 40% after 1 month at 40°C. Figure 28B shows acidic variant formation at pH 5.5-6.0, showing that the % acidic variant remained generally below 30.0% after 3 months at 25°C. Figure 28C shows acidic variant formation at all pHs, showing that the % acidic variant remained generally below 25% after 6 months at 5°C. Figures 29A-C show the effect of pH between pH 5.5-6.5 on basic variant formation in histidine / sucrose formulations containing 20 mM histidine, 270 mM sucrose, and 0.01% polysorbate 20. Figure 29A shows basic variant formation at pH 5.5-6.0, showing that the % basic variant remained mostly between 10%-20% after 1 month at 40°C. Figure 29B shows basic variant formation at pH 5.5-6.0, showing that the % basic variant remained generally between 20%-30% after 3 months at 25°C. Figure 29C shows basic variant formation at all pHs, showing that the % basic variant remained generally between 25%-30% after 6 months at 5°C.

[0191] Figures 30A-C show the effect of pH between pH 5.5-6.5 on basic variant formation in a histidine / arginine formulation containing 20 mM histidine, 150 mM arginine, and 0.01% polysorbate 20. Figure 30A shows basic variant formation at pH 5.5-6.0, showing that the % basic variant remained mostly between 10%-20% after 1 month at 40°C. Figure 30B shows basic variant formation at pH 5.5-6.0, showing that the % basic variant remained generally between 15%-25% after 3 months at 25°C. Figure 30C shows basic variant formation at all pHs, showing that the % basic variant remained generally between 25%-30% after 6 months at 5°C.

[0192] Both the histidine / arginine and histidine / sucrose formulations maintained acceptable stability between pH 5.5 and pH 6.5. Overall, the histidine / sucrose formulation was more stable than the histidine / arginine between pH 5.5 and 6.0.

[0193] Based on the data regarding the formulation stability at 25° C. for 3 months and at 5° C. for 6 months, as well as data from other formulation studies in the Examples above, a liquid formulation was developed containing 20 mg / mL anti-FGFR2-IIIb antibody, 20 mM L-histidine, 270 mM sucrose, and 0.01% polysorbate 20 at a pH of 6.0. A second formulation was also developed containing 20 mg / mL anti-FGFR2-IIIb antibody, 20 mM L-histidine, 150 mM L-arginine, 0.01% polysorbate 20 at a pH of 5.7.

[0194] The present invention should not be limited in scope by the specific embodiments described herein. Indeed, various modifications provided herein in addition to those described will become apparent to those skilled in the art from the foregoing description and accompanying drawings. Such modifications are intended to fall within the scope of the appended claims.

[0195] All references (e.g., publications or patents or patent applications) cited in this specification are incorporated by reference in their entirety and for all purposes to the same extent as if each individual reference (e.g., publication or patent or patent application) was specifically and individually indicated to be incorporated by reference in its entirety for all purposes.

[0196] Other embodiments are within the scope of the following claims.

[0197] Array Table The following sequence listing provides certain sequences discussed herein. Unless otherwise indicated, all polypeptide and antibody sequences are shown without leader sequences. TIFF2024138284000013.tif253165TIFF2024138284000014.tif253161TIFF2024138284000015.tif253163TIFF2024138284000016.tif16169

Claims

1. A liquid pharmaceutical formulation, wherein the pharmaceutical formulation is not lyophilized; i) 10-30 mg / mL of an anti-fibroblast growth factor receptor 2 (FGFR2) antibody; ii) 5-40 mM buffer selected from one or more of histidine, citrate, or phosphate; iii) 130-170 mM arginine; iv) 0.002% to 0.1% polysorbate 20 or polysorbate 80; Including, The formulation has a pH of 5.0 to 6.5, and the anti-FGFR2 antibody is a) an antibody comprising a heavy chain amino acid sequence of SEQ ID NO:2 and a light chain amino acid sequence of SEQ ID NO:3; b) an antibody comprising a heavy chain (HC) hypervariable region 1 (HVR1) comprising the amino acid sequence of SEQ ID NO:6, a HC HVR2 comprising the amino acid sequence of SEQ ID NO:7, and a HC HVR3 comprising the amino acid sequence of SEQ ID NO:8, and a light chain (LC) HVR1 comprising the amino acid sequence of SEQ ID NO:9, a LC HVR2 comprising the amino acid sequence of SEQ ID NO:10, and a LC HVR3 comprising the amino acid sequence of SEQ ID NO:11; or c) An antibody comprising the heavy chain variable region amino acid sequence of SEQ ID NO: 4 and the light chain variable region amino acid sequence of SEQ ID NO:

5.

1. A liquid pharmaceutical formulation comprising:

2. 2. The liquid pharmaceutical formulation of claim 1, wherein the formulation comprises 10 mg / mL, 11 mg / mL, 12 mg / mL, 13 mg / mL, 14 mg / mL, 15 mg / mL, 16 mg / mL, 17 mg / mL, 18 mg / mL, 19 mg / mL, 20 mg / mL, 21 mg / mL, 22 mg / mL, 23 mg / mL, 24 mg / mL, or 25 mg / mL of the anti-FGFR2 antibody.

3. 3. The liquid pharmaceutical formulation of any one of claims 1-2, wherein the formulation comprises 10 mM, 15 mM, 16 mM, 17 mM, 18 mM, 19 mM, 20 mM, 21 mM, 22 mM, 23 mM, 24 mM, 25 mM, 26 mM, 27 mM, 28 mM, 29 mM, or 30 mM of the buffer.

4. The liquid pharmaceutical formulation of claim 1, further comprising a histidine buffer.

5. 5. The liquid pharmaceutical formulation of any one of claims 1 to 4, wherein the formulation comprises 130 mM, 135 mM, 140 mM, 145 mM, 150 mM, 155 mM, 160 mM, 165 mM, or 170 mM arginine.

6. The liquid pharmaceutical formulation of any one of claims 1 to 5, wherein the formulation is sucrose-free.

7. 7. The liquid pharmaceutical formulation of any one of claims 1 to 6, wherein the pH of the formulation is 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, or 6.

5.

8. 8. The liquid pharmaceutical formulation of any one of claims 1 to 7, wherein the formulation comprises 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 or 80.

9. 9. The liquid pharmaceutical formulation of claim 8, wherein the formulation comprises polysorbate 20.

10. The formulation comprises: i) 10 to 25 mg / mL of the anti-FGFR2 antibody; ii) 10-30 mM histidine buffer; iii) 140-160 mM arginine; iv) 0.005% to 0.05% polysorbate 20; 10. The liquid pharmaceutical formulation of claim 1, wherein the formulation has a pH of 5.6 to 5.

8.

11. The formulation comprises: i) 10 to 25 mg / mL of the anti-FGFR2 antibody; ii) 20 mM histidine buffer; iii) 150 mM arginine; iv) 0.01% polysorbate 20, 11. The liquid pharmaceutical formulation of claim 10, wherein the formulation has a pH of 5.

7.

12. The formulation comprises: i) 20 mg / mL of the anti-FGFR2 antibody; ii) 20 mM L-histidine buffer; iii) 150 mM L-arginine; iv) 0.01% polysorbate 20, 10. The liquid pharmaceutical formulation of claim 1, wherein the formulation has a pH of 5.

7.

13. 2. The liquid pharmaceutical formulation of claim 1, wherein the formulation consists essentially of the anti-FGFR2 antibody, histidine buffer, arginine, and polysorbate 20.

14. The liquid pharmaceutical formulation of any one of claims 1 to 13, wherein the anti-FGFR2 antibody is afucosylated.

15. The liquid pharmaceutical formulation of any one of claims 1 to 14, wherein the anti-FGFR2b antibody comprises a heavy chain amino acid sequence of SEQ ID NO:2 and a light chain amino acid sequence of SEQ ID NO:

3.

16. The liquid pharmaceutical formulation of any one of claims 1 to 15, for treating a solid tumor in a patient in need thereof.

17. 17. The liquid pharmaceutical formulation of claim 16, wherein the formulation is administered intravenously to the patient.

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