Risankizumab composition

Abstract

The present disclosure relates, in part, to risankizumab compositions having reduced levels of the hitchhiker protein PLA2, poloxamer 188, and / or reduced immunogenicity.

Description

[Technical Field] 【0001】 CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Application No. 63 / 352,459, filed June 15, 2022, U.S. Provisional Application No. 63 / 455,495, filed March 29, 2023, U.S. Provisional Application No. 63 / 444,178, filed February 8, 2023, and U.S. Provisional Application No. 63 / 444,182, filed February 8, 2023, the contents of each of which are incorporated herein by reference in their entirety. 【0002】 Sequence table XML reference This application contains a Sequence Listing that has been submitted electronically in XML format. This Sequence Listing is incorporated herein by reference. The XML file, created on June 13, 2023, is named AVR-05525_SL.xml and is 16,072 bytes in size. [Background technology] 【0003】 Risankizumab (approved by the Food and Drug Administration (FDA) in the United States as risankizumab-rzaa and sold under the trade name SKYRIZI®) is a humanized immunoglobulin G1 (IgG1) monoclonal antibody directed against the p19 subunit of IL-23. Risankizumab's binding to IL-23 p19 inhibits IL-23's ability to induce and maintain T helper (Th)17 type cells, innate lymphoid cells, gamma delta T cells, and natural killer (NK) cells, which are involved in tissue inflammation, destruction, and abnormal tissue repair. Risankizumab is particularly effective in treating autoimmune and inflammatory diseases, such as psoriasis. Clinical trials have demonstrated excellent safety and efficacy in the treatment of, for example, plaque psoriasis and psoriatic arthritis. 【0004】 Risankizumab can be formulated at different concentrations for subcutaneous injection. For example, risankizumab formulations at concentrations of 60 mg / mL, 90 mg / mL, and 150 mg / mL have been approved by the FDA. Various risankizumab formulations are described in International Applications PCT / US2013 / 038109 and PCT / IB2020 / 058347, the contents of which are incorporated herein by reference in their entireties. 【0005】 The above-mentioned commercial formulations contain the surfactant polysorbate 20 (PS20). It has been proposed that trace amounts of hitchhiker protein contaminants in preparations of certain recombinantly produced biopharmaceuticals can cause hydrolysis of polysorbate 20, leading to particle formation (therapeutic protein and / or free fatty acid aggregates), and thus reducing shelf life (Khan et al. (2015) European Journal of Pharmaceutics and Biopharmaceutics 97:60-67). 【0006】 One important factor contributing to the presence of hitchhiker protein impurities in recombinant therapeutic monoclonal antibody preparations is their association with the produced monoclonal antibodies (mAbs) (Nogal et al. (2012) Biotechnol.Prog. 28:454-458). It has been reported that mAbs can preferentially bind to selected hitchhiker proteins (HPs), and the degree of interaction and / or identity of associated HPs can vary depending on the mAb (Nogal et al. (2012) Biotechnol.Prog. 28:454-458). It has also been shown that the hitchhiker protein content of mAbs in Protein A (PrA) eluates is specific to a particular antibody (Zhang et al. (2016) Biotechnol. Prog. 32:708-717), and that the primary sequence of a mAb can influence the binding and resulting co-purification of specific hitchhiker proteins (Bee et al. (2016) Biotechnol. Prog. 00:1-6). Another factor is that certain hitchhiker proteins have similar physicochemical properties to the specific mAb being purified, leading to their co-purification with the mAb. Because different hitchhiker proteins co-purify with different recombinant antibodies, it is impossible to predict before an experiment whether hitchhiker problems will arise during the production of a new antibody, let alone whether hitchhiker protein problems will arise. 【0007】 Furthermore, a major problem with protein-based therapeutics is their immunogenicity, i.e., their tendency to elicit unwanted immune responses against themselves, resulting in so-called "anti-drug antibodies" or "ADAs." 【0008】 To date, there remains a need to develop risankizumab compositions with reduced levels of hitchhiker protein impurities and improved properties, such as reduced immunogenicity. [Prior art documents] [Non-patent literature] 【0009】 [Non-Patent Document 1] Khan et al. (2015) European Journal of Pharmaceutics and Biopharmaceutics 97:60-67 [Non-patent document 2] Nogal et al. (2012) Biotechnol.Prog. 28:454-458 [Non-patent document 3] Zhang et al. (2016) Biotechnol.Prog. 32:708-717 [Non-patent document 4] Bee et al. (2016) Biotechnol.Prog. 00:1-6 Summary of the Invention [Means for solving the problem] 【0010】 The present disclosure is based, in part, on the discovery of a specific hitchhiker protein, phospholipase A2 (PLA2), that copurifies with risankizumab. The presence of this PLA2 adversely affects the stability of polysorbates (e.g., polysorbate 20 and / or polysorbate 80) in risankizumab liquid pharmaceutical formulations. Reducing the PLA2 concentration in the formulation beneficially increases the formulation's long-term stability (e.g., reducing particle formation and extending the shelf life of risankizumab drug products). Increased stability of risankizumab formulations can also be achieved by using poloxamer 188 (P188) instead of PS20 or PS80. The present disclosure also describes novel risankizumab compositions with reduced levels and improved purity of risankizumab species modified with high-mannose N-glycans (e.g., M5, M6, and / or M7). These risankizumab compositions exhibit reduced immunogenicity in human subjects. 【0011】 Accordingly, in one aspect, the present disclosure relates to a liquid composition comprising: (1) risankizumab; and (2) PLA2 in an amount that is less than about 250 pg per mg of risankizumab. 【0012】 In another aspect, the disclosure relates to a composition comprising risankizumab, wherein the composition has one or more of the following characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high mannose N-glycans, and / or (b) the incidence of treatment-emergent anti-drug antibodies (ADA) in humans is less than about 4.7% following administration of a single 150 mg subcutaneous dose to said humans. 【0013】 In yet another aspect, the disclosure relates to a composition comprising (1) risankizumab and (2) poloxamer 188 (P188), wherein the composition does not include polysorbate 20 (PS20) and / or polysorbate 80 (PS80). [Brief explanation of the drawings] 【0014】 [Figure 1] The number of affinity-purified low molecular weight (LMW) hitchhiker proteins (HPs) in DP1 and DP2, total HPs, and LMW HPs and total HPs are shown. [Figure 2] Western blot probed with anti-PLA2G15 antibody is shown: lane 1, MW standard; lane 2, 1 ng of PLA2G15 (MW: 47 kDa); lane 3, 0.1 ng of PLA2G15; lane 6, DP1; lane 7, DP2_#1; lane 8, DP2_#2; lane 9, DP3, and lane 10, DP4. [Figure 3A] 1 shows the stability of PS20 at 5° C. in samples made from various control and knockout cell lines as measured by CAD assay. [Figure 3B] Figure 1 shows the stability of PS20 at 25°C in samples made from various control and knockout cell lines as measured by CAD assay. #5 Placebo: PS20 control. #6 BDS control: DP4 BDS. [Figure 3C]Shows the stability of PS20 at 5°C in samples made from various control and knockout cell lines as measured by FFA assay. #5: Placebo: PS20 control. #6: BDS control: DP4 BDS. [Figure 3D] Shows the stability of PS20 at 25°C in samples made from various control and knockout cell lines as measured by FFA assay. #5: Placebo: PS20 control. #6: BDS control: DP4 BDS. [Figure 4A] Overlap of PS20 subspecies chromatograms for sample C1 injection 1 (DP2 after 30 months at 2–8 °C), sample A1 injection 2 (DP3 spiked with 1 μg / mL of PLA2G15 after approximately 9 hours of incubation at room temperature and 25 °C), and sample D1 injection 1 (unspiked DP3 material, no significant PS20 degradation) are shown. [Figure 4B] Figure 2 shows the overlay of PS20 subspecies chromatograms of PS20 degradation in DP4 solution at different PLA2G15 spike levels (more sample information can be found in Table 20) after 4 days of incubation at 25°C. Group 8 was the DP2 control sample. The small difference near 42 minutes in Group 8 may be caused by eluates from the syringe filter used in this in-lab loading for this group, as this peak was not observed in the previous data. [Figure 5] Overlapping PS20 subspecies chromatograms for sample E3 injection 2 (DP3, no significant PS20 degradation), sample A3 injection 6 (DP3 spiked with 5 μg / mL PLBL2 after approximately 30 hours of incubation at 25°C; total incubation time after spiking was approximately 20 hours at 2-8°C + 26 hours at 25°C), and sample D3 injection 2 (PS20 in DP2 after approximately 30 months of storage at 2-8°C) are shown. [Figure 6A]Overlap of PS20 subspecies chromatograms for sample E3 injection 2 (DP3, no significant PS20 degradation) and sample B3 injection 6 (DP3 spiked with 5 μg / mL CES1 after approximately 30 hours of incubation at 25 °C; total incubation time after spiking was approximately 20 hours at 2–8 °C + 27 hours at 25 °C). [Figure 6B] Overlap of PS20 subspecies chromatograms for sample D3 injection 2 (PS20 degradation of DP2 after approximately 30 months of storage at 2–8°C) and sample B3 injection 6 (DP3 spiked with 5 μg / mL CES1 after approximately 30 hours of incubation at 25°C; total incubation time after spiking was approximately 20 hours at 2–8°C + 27 hours at 25°C). [Figure 7] Overlap of PS20 subspecies chromatograms for sample E3 injection 2 (DP3, no significant PS20 degradation) and sample C3 injection 6 (DP3 spiked with 5 μg / mL SIAE after approximately 30 hours of incubation at 25 °C; total incubation time after spiking was approximately 20 hours at 2–8 °C + 28 hours at 25 °C). [Figure 8] Overlapping PS20 subspecies chromatograms for sample D4 injection 2 (DP3, no significant PS20 degradation), sample A4 injection 7 (DP3 spiked with 5 μg / mL PRDX6 after approximately 30 hours of incubation at 25°C; total incubation time after spiking was approximately 27 hours at 25°C), and sample C4 injection 2 (PS20 in DP2 after approximately 30 months of storage at 2–8°C) are shown. [Figure 9] Overlapping PS20 subspecies chromatograms for sample D4 injection 2 (DP3, no significant PS20 degradation), sample B4 injection 7 (DP3 spiked with 5 μg / mL PLA2G7 after approximately 30 hours of incubation at 25 °C; total incubation time after spiking was approximately 28 hours at 25 °C), and sample C4 injection 2 (PS20 in DP2 after approximately 30 months of storage at 2–8 °C) are shown. [Figure 10]Overlapped PS20 variant chromatograms for samples H7-9, H7-8, and H7-7 are shown. H7-9: DP2 (held at -80°C); H7-8: DP2 (held at room temperature for 2 weeks); H7-7: DP2 spiked with 0.9 μg / mL fosinopril (held at room temperature for 2 weeks). Signals were normalized to correct for concentration changes due to spiking (normalized to the peak at 37 minutes, which is stable in the DP2 material based on historical data). [Figure 11] Overlapped PS20 variant chromatograms for samples H7-9, H7-8, and H7-6 are shown. H7-9: DP2 (held at -80°C); H7-8: DP2 (held at room temperature for 2 weeks); H7-6: DP2 spiked with 3.8 μg / mL fosinopril (held at room temperature for 2 weeks). Signals were normalized to correct for concentration changes due to spiking (normalized to the peak at 37 minutes, which is stable in the DP2 material based on historical data). [Figure 12] Overlapped PS20 variant chromatograms for samples H7-9, H7-8, and H7-3 are shown. H7-9: DP2 (held at -80°C); H7-8: DP2 (held at room temperature for 2 weeks); H7-3: DP2 spiked with 27.8 μg / mL fosinopril (held at room temperature for 2 weeks). Signals were normalized to correct for concentration changes due to spiking (normalized to the peak at 37 minutes, which is stable in the DP2 material based on historical data). The higher background in the spiked sample around 39-42 minutes is likely due to coelution of fosinopril. [Figure 13] Overlap of PS20 variant chromatograms for samples A6 and C6 are shown. A6:DP2 material (held at -80°C, diluted 1:1 with water before testing); C6:DP2 material (spiked with 930 μg / mL fosinopril; held at room temperature for 2 weeks; diluted 1:1 with water before testing). The signal from C6 was normalized for better comparison with A5. [Figure 14] A general overview of the newly developed purification process for risankizumab drug substance (referred to herein as Process 4 Development) is shown. [Figure 15A] 1 shows the stability of PS20 in DP2(PS20) and DP3(PS20) as measured by CAD assay at 5° C. [Figure 15B] 1 shows the stability of PS20 in DP2(PS20) and DP3(PS20) as measured by CAD assay at 25° C. [Figure 15C] 1 shows the stability of PS20 in DP2(PS20) and DP3(PS20) as measured by CAD assay at 40° C. [Figure 16A] 1 shows the stability of PS20 in DP2(PS20) and DP3(PS20) as measured by FFA assay at 5° C. [Figure 16B] 1 shows the stability of PS20 in DP2(PS20) and DP3(PS20) as measured by FFA assay at 25° C. [Figure 17A] 1 shows the stability of PS20 in DP2(PS20) and DP4(PS20) as measured by CAD assay at 5° C. [Figure 17B] 1 shows the stability of PS20 in DP2(PS20) and DP4(PS20) as measured by CAD assay at 25° C. [Figure 17C] 1 shows the stability of PS20 in DP2(PS20) and DP4(PS20) as measured by CAD assay at 40° C. [Figure 18A] 1 shows the stability of PS20 in DP2(PS20) and DP4(PS20) as measured by FFA assay at 5° C. [Figure 18B] 1 shows the stability of PS20 in DP2(PS20) and DP4(PS20) as measured by FFA assay at 5° C. [Figure 18C] 1 shows the stability of PS20 in DP2(PS20) and DP4(PS20) as measured by FFA assay at 25° C. [Figure 18D] 1 shows the stability of PS20 in DP2(PS20) and DP4(PS20) as measured by FFA assay at 25° C. [Figure 19A] 1 shows the stability of PS80 in DP2(PS80), DP3(PS80), and DP4(PS80) as measured by CAD assay at 5° C. [Figure 19B]1 shows the stability of PS80 in DP2(PS80), DP3(PS80) and DP4(PS80) as measured by CAD assay at 25° C. [Figure 19C] 1 shows the stability of PS80 in DP2(PS80), DP3(PS80) and DP4(PS80) as measured by CAD assay at 40° C. [Figure 20A] 1 shows the stability of PS80 in DP2(PS80), DP3(PS80), and DP4(PS80) as measured by FFA assay at 5° C. [Figure 20B] 1 shows the stability of PS80 in DP2(PS80), DP3(PS80), and DP4(PS80) as measured by FFA assay at 5° C. [Figure 20C] 1 shows the stability of PS80 in DP2(PS80), DP3(PS80) and DP4(PS80) as measured by FFA assay at 25° C. [Figure 20D] 1 shows the stability of PS80 in DP2(PS80), DP3(PS80) and DP4(PS80) as measured by FFA assay at 25° C. [Figure 20E] 1 shows the stability of PS80 in DP2(PS80), DP3(PS80) and DP4(PS80) as measured by FFA assay at 40° C. [Figure 20F] 1 shows the stability of PS80 in DP2(PS80), DP3(PS80) and DP4(PS80) as measured by FFA assay at 40° C. [Figure 21A] 2-AB and HILIC-FL chromatograms of a Process 4 drug substance (DS) batch and Process 1 reference standard DS1-RS2 are shown. Process 4 batch DS4-001 and Process 1 reference standard DS1-RS2 were analyzed in parallel. Results for the reference standards accompanying the other three Process 4 DS batches are not shown. As expected, slight differences in retention times from different runs were observed. Assay performance (relative peak quantification) is unaffected. [Figure 21B] FIG. 21B is an enlarged view of FIG. 21A. [Figure 22]RapiFluor and HILIC-FL chromatograms of DS batches of processes 1, 2, and 4 are shown. [Figure 23A] 1 shows the relative distribution of UP-SEC monomer results for DS batches of risankizumab processes 1, 2, and 4. [Figure 23B] 1 shows the relative distribution of UP-SEC HMW results for DS batches of risankizumab processes 1, 2, and 4. [Figure 23C] Figure 1 shows UP-SEC results for risankizumab Process 4 DS batch DS4-005 and Process 1 reference standard DS1-RS2. [Figure 23D] FIG. 23D is an enlarged view of FIG. 23C. [Figure 24A] 1 shows the relative distribution of CGE-NR main peak results for DS batches of risankizumab processes 1, 2, and 4. [Figure 24B] 1 shows the relative distribution of CGE-NR LMW results for DS batches of risankizumab processes 1, 2, and 4. [Figure 24C] Figure 1 shows CGE-NR results for risankizumab Process 4 DS batch DS4-005 and Process 1 reference standard DS1-RS2. [Figure 24D] An enlarged view of Figure 24C is shown. [Figure 25] The levels of P188 and PS20 in the DS of DP2 are shown. [Figure 26] P188 and PS20 levels in DS of DP3 are shown. DETAILED DESCRIPTION OF THE INVENTION 【0015】 In some embodiments, the present disclosure is based, in part, on the discovery of a specific hitchhiker protein, PLA2. The presence of this PLA2 adversely affects the stability of polysorbates (e.g., PS20 and / or PS80) in risankizumab liquid pharmaceutical formulations, and reducing PLA2 from the formulation beneficially increases the stability of the formulation (e.g., reducing particle formation, extending the shelf life of risankizumab drug products, etc.). It has also been found that increased stability of risankizumab formulations can also be achieved by using poloxamer 188 in place of P20 or P80. 【0016】 The initial pharmaceutical formulation developed for risankizumab had a concentration of 90 mg / ml. A 150 mg / ml formulation was subsequently approved by the U.S. FDA, allowing for a single subcutaneous injection of the full 150 mg therapeutic dose. Both the commercially available 75 mg / 0.83 ml (90 mg / mL) and 150 mg / ml risankizumab formulations are disclosed in the FDA-approved drug labeling and the full prescribing information for SKYRIZI® (risankizumab-rzaa) revised in December 2022, the contents of each of which are incorporated herein by reference in their entirety. Both FDA-approved risankizumab formulations contain highly purified, recombinantly produced risankizumab active pharmaceutical ingredient (API). However, when a 150 mg / ml risankizumab formulation was diluted to explore the feasibility of developing specific product presentations, such as those used with on-body devices, unacceptable levels of particles comprising risankizumab and / or free fatty acid aggregates were formed under certain storage conditions. 【0017】 In one embodiment of the present invention, this unexpected problem is believed to be due to the persistence of trace amounts of hitchhiker proteins co-purified with otherwise highly pure risankizumab API purified by state-of-the-art orthogonal column chromatography processes. Because the identity of hitchhiker proteins co-purified with monoclonal antibodies (mAbs) varies depending on the mAb, it is impossible to predict prior to an experiment whether hitchhiker protein issues will arise during the production of a new antibody, let alone whether hitchhiker protein issues will arise at all. 【0018】 In some embodiments, the present disclosure identifies PLA2 as a particular problematic hitchhiker protein co-purified with risankizumab. It is demonstrated herein that PLA2 co-purified with risankizumab causes degradation of the surfactant polysorbate 20 (PS20), resulting in particle formation in the risankizumab product. An optimized purification process has been developed specifically to reduce the level of PLA2 co-purified with risankizumab. Thus, in some embodiments, the present disclosure provides risankizumab liquid compositions with reduced levels of PLA2 and improved stability and shelf life. 【0019】 In some aspects, the present disclosure relates to novel risankizumab compositions having reduced levels of risankizumab species modified with high mannose N-glycans (e.g., M5, M6, and / or M7) that have reduced immunogenicity. 【0020】 In some aspects, the present disclosure relates to risankizumab compositions having reduced levels of risankizumab species having high mannose N-glycans (M5, M6, and / or M7) and reduced immunogenicity. Reduced immunogenicity (e.g., reduced incidence of therapeutically emerged anti-drug antibodies following administration of a single 150 mg subcutaneous dose of the liquid composition to humans) also indicates improved product quality of the risankizumab compositions described herein. 【0021】 As disclosed herein, the present disclosure relates to the following embodiments: 【0022】 Embodiment 1. A liquid composition comprising: (1) risankizumab or an anti-IL23 monoclonal antibody comprising two light chains having the amino acid sequence of SEQ ID NO:9 and two heavy chains having the amino acid sequence of SEQ ID NO:10; and (2) phospholipase A2 (PLA2) in an amount that is less than about 250 pg per mg of risankizumab. 【0023】 Embodiment 2. The liquid composition of embodiment 1, comprising about 60 mg / ml to about 150 mg / ml of risankizumab. 【0024】 Embodiment 3. The liquid composition of embodiment 1 or 2, wherein the PLA2 is PLA2G15. 【0025】 Embodiment 4. The level of PLA2 is less than about 240 pg, less than about 220 pg, less than about 200 pg, less than about 180 pg, less than about 160 pg, less than about 140 pg, less than about 120 pg, less than about 100 pg, less than about 90 pg, less than about 80 pg, less than about 70 pg, less than about 60 pg, less than about 50 pg, less than about 40 pg, less than about 30 pg, less than about 25 pg, or less than about 30 pg per mg of risankizumab. 4. The liquid composition of any one of embodiments 1-3, wherein the α-tocopherol concentration is less than about 20 pg, less than about 15 pg, less than about 10 pg, less than about 9 pg, less than about 8 pg, less than about 7 pg, less than about 6 pg, less than about 5 pg, less than about 4.4 pg, less than about 3 pg, less than about 2 pg, less than about 1 pg, less than about 0.5 pg, less than about 0.1 pg, less than about 0.05 pg, or less than about 0.01 pg. 【0026】 Embodiment 5. The level of PLA2 is greater than about 240 pg, greater than about 220 pg, greater than about 200 pg, greater than about 180 pg, greater than about 160 pg, greater than about 140 pg, greater than about 120 pg, greater than about 100 pg, greater than about 90 pg, greater than about 80 pg, greater than about 70 pg, greater than about 60 pg, greater than about 50 pg, greater than about 40 pg, greater than about 30 pg, greater than about 4. The liquid composition of any one of embodiments 1-3, wherein the soluble ... 【0027】 Embodiment 6. The liquid composition of any one of embodiments 1 to 3, wherein the level of PLA2 is about 200 to about 249, about 160 to about 200, about 120 to about 160, about 100 to about 120, about 80 to about 100, about 60 to about 80, about 40 to about 60, about 25 to about 40, about 10 to about 25, about 5 to about 10, about 4 to about 10, about 1 to about 5, about 1 to about 4, about 1 to about 3, about 1 to about 2, about 0.5 to about 1, about 0.1 to about 0.5, about 0.05 to about 0.1, about 0.01 to about 0.5 pg, or about 70 to about 240 pg per mg of risankizumab. 【0028】 Embodiment 7. The liquid composition of any one of embodiments 1 to 3, wherein the level of PLA2 is about 240, about 220, about 200, about 180, about 160, about 140, about 120, about 100, about 90, about 80, about 70, about 60, about 50, about 40, about 30, about 25, about 20, about 15, about 10, about 9, about 8, about 7, about 6, about 5, about 4.4, about 3, about 2, about 1, about 0.5, about 0.1, about 0.05, or about 0.01 pg / mg of risankizumab. 【0029】 Embodiment 8. The liquid composition of any one of embodiments 1 to 7, wherein the level of PLA2 is determined by ELISA. 【0030】 Embodiment 9. The liquid composition of any one of embodiments 1 to 8, wherein the risankizumab is produced in a CHO cell line. 【0031】 Embodiment 10. The liquid composition of any one of embodiments 1 to 9, further comprising one or more of a surfactant, a polyol, and a buffering agent. 【0032】 Embodiment 11. The liquid composition of embodiment 10, wherein the polyol is selected from the group consisting of trehalose, mannitol, sucrose, and sorbitol. 【0033】 Embodiment 12. The liquid composition of embodiment 11, wherein the polyol is trehalose. 【0034】 Embodiment 13. The liquid composition of embodiment 12, wherein the trehalose is present in an amount of about 150 to about 220 mM. 【0035】 Embodiment 14. The liquid composition of embodiment 13, wherein the trehalose is present in an amount of about 185 mM. 【0036】 Embodiment 15. The liquid composition of any one of embodiments 10 to 14, wherein the buffer is selected from the group consisting of acetate buffer, histidine buffer, citrate buffer, phosphate buffer, glycine buffer, and arginine buffer. 【0037】 Embodiment 16. The liquid composition of embodiment 15, wherein the buffer is an acetate buffer. 【0038】 Embodiment 17. The liquid composition of embodiment 16, wherein the acetate buffer is present in an amount of about 5 to about 50 mM. 【0039】 Embodiment 18. The liquid composition of embodiment 17, wherein the acetate buffer is present in an amount of about 10 mM. 【0040】 Embodiment 19. The liquid composition of any one of embodiments 10 to 18, wherein the surfactant is selected from the group consisting of polysorbate 20 (PS20), polysorbate 80 (PS80), polysorbate 40 (PS40), polysorbate 60 (PS60), polysorbate 65 (PS65), and poloxamer 188. 【0041】 Embodiment 20. The liquid composition of embodiment 19, wherein the surfactant is PS20. 【0042】 Embodiment 21. The liquid composition of embodiment 20, wherein the PS20 is present in an amount of about 0.2 mg / mL. 【0043】 Embodiment 22. The liquid composition of embodiment 21 comprising 150 mg / ml risankizumab, 185 mM trehalose, 10 mM acetate, and 0.20 mg / mL polysorbate 20, wherein the pH is about 5.7. 【0044】 Embodiment 23. The liquid composition of embodiment 21 comprising 150 mg / ml risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, and water for injection, wherein the pH is about 5.7. 【0045】 Embodiment 2: The liquid composition of any one of embodiments 20-23, wherein at least 80% of the initial concentration of PS20 is present in the composition after storage at 4.5°C for 6 months. 【0046】 Embodiment 25. The liquid composition of any one of embodiments 20-23, wherein at least 70% of the initial concentration of PS20 is present in the composition after 24 months of storage at 5°C. 【0047】 Embodiment 26. The liquid composition of any one of embodiments 20-23, wherein at least 60% of the initial concentration of PS20 is present in the composition after storage at 25°C for 6 months. 【0048】 Embodiment 27. The liquid composition of any one of embodiments 20-23, wherein at least 40% of the initial concentration of PS20 is present in the composition after storage at 40°C for 6 months. 【0049】 Embodiment 2: The liquid composition of any one of embodiments 20-23, wherein after storage at 8.5°C for 6 months, the total concentration of free fatty acids (FFA) present in the composition increases by no more than 1.5-fold. 【0050】 Embodiment 29. The liquid composition of any one of embodiments 20-23, wherein after 6 months of storage at 5°C, the total concentration of FFAs present in the composition is 20 nmol / ml or less. 【0051】 Embodiment 30. The liquid composition of any one of embodiments 20-23, wherein after 6 months of storage at 25°C, the total concentration of FFAs present in said composition is 3.2 times or less. 【0052】 Embodiment 31. The liquid composition of any one of embodiments 20-23, wherein after 6 months of storage at 25°C, the total concentration of FFAs present in the composition is 25 nmol / ml or less. 【0053】 Embodiment 32. The liquid composition of any one of embodiments 20-23, wherein the total concentration of FFAs present in the composition is no more than 3-fold after storage at 40°C for 6 months. 【0054】 Embodiment 33. The liquid composition of any one of embodiments 20-23, wherein after 6 months of storage at 40°C, the total concentration of FFAs present in said composition is 35 nmol / ml or less. 【0055】 Embodiment 34. The liquid composition of embodiment 19, wherein the surfactant is PS80. 【0056】 Embodiment 35. The liquid composition of embodiment 34, wherein at least 80% of the initial concentration of PS80 is present in the composition after 6 months of storage at 5°C. 【0057】 Embodiment 36. The liquid composition of embodiment 34, wherein after 6 months of storage at 25°C, at least 60% of the initial concentration of PS80 is present in the composition. 【0058】 Embodiment 37. The liquid composition of embodiment 34, wherein after 6 months of storage at 40°C, at least 60% of the initial concentration of PS80 is present in the composition. 【0059】 Embodiment 38. The liquid composition of embodiment 34, wherein the total concentration of FFAs present in the composition is 8-fold or less after 6 months of storage at 5°C. 【0060】 Embodiment 39. The liquid composition of embodiment 34, wherein after 6 months of storage at 5° C., the total concentration of FFAs present in the composition is 40 nmol / ml or less. 【0061】 Embodiment 40. The liquid composition of embodiment 34, wherein the total concentration of FFAs present in the composition is no more than 12-fold after 6 months of storage at 25°C. 【0062】 Embodiment 41. The liquid composition of embodiment 34, wherein after 6 months of storage at 25°C, the total concentration of FFAs present in the composition is 60 nmol / ml or less. 【0063】 Embodiment 42. The liquid composition of embodiment 34, wherein after 6 months of storage at 40°C, the total concentration of FFAs present in the composition increases by no more than 2.5-fold. 【0064】 Embodiment 43. The liquid composition of embodiment 34, wherein after 6 months of storage at 40°C, the total concentration of FFAs present in the composition is 15 nmol / ml or less. 【0065】 Embodiment 44. The liquid composition of any one of embodiments 24 to 43, wherein the PS20 or the PS80 is measured using HPLC-CAD. 【0066】 Embodiment 45. The liquid composition of any one of embodiments 24 to 43, wherein the FFAs are measured using an LC-FFA assay. 【0067】 Embodiment 46. The liquid composition of any one of embodiments 1 to 45, wherein no visible or shiny particles are observed at 4°C for 24 months. 【0068】 Embodiment 47. The liquid composition of any one of embodiments 1 to 46, wherein the liquid composition is packaged in a vial, a pre-filled syringe, or an on-body device. 【0069】 Embodiment 48. The liquid composition of any one of embodiments 1 to 47, wherein the liquid composition is a pharmaceutical composition and is suitable for subcutaneous injection. 【0070】 Embodiment 49. The liquid composition of any one of embodiments 1 to 47, wherein the liquid composition is a pharmaceutical composition and is suitable for intravenous injection. 【0071】 Embodiment 50. A method of treating an immunological disease with a composition according to any one of embodiments 1 to 49. 【0072】 Embodiment 51. The method of claim 51, wherein the composition comprises risankizumab, wherein the composition has one or more of the following characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans; (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by ultra-performance size-exclusion chromatography (UP-SEC); (c) more than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as a low molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR); and / or (d) the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% after administration of a single subcutaneous dose of the composition to a human. 【0073】 Embodiment 52. The composition of embodiment 51, comprising about 60 mg / ml to about 150 mg / ml of risankizumab. 【0074】 Embodiment 53. The composition of embodiment 51 or 52, wherein the pharmaceutical composition has at least the characteristic (a) that less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans. 【0075】 Embodiment 54. The composition of embodiment 53, wherein the high mannose N-glycans comprise one or more high mannose N-glycans selected from mannose 5 N-glycan (M5), mannose 6 N-glycan (M6), and mannose 7 N-glycan (M7). 【0076】 Embodiment 55. The composition of embodiment 54, wherein the high mannose N-glycans are M5, M6, and M7. 【0077】 Embodiment 56. The composition of any one of embodiments 51-55, wherein the level of risankizumab containing the high mannose N-glycans is less than 5.3%, less than about 5.2%, less than about 5.1%, less than about 5.0%, less than about 4.9%, less than about 4.8%, less than about 4.7%, less than about 4.6%, less than about 4.5%, less than about 4.4%, less than about 4.3%, less than about 4.2%, less than about 4.1%, less than about 4.0%, less than about 3.9%, less than about 3.8%, or less than about 3.7% of total risankizumab species with N-glycosylation. 【0078】 Embodiment 57. The composition of any one of embodiments 51-56, wherein the level of risankizumab containing the high mannose N-glycans is greater than about 5.3%, greater than about 5.2%, greater than about 5.1%, greater than about 5.0%, greater than about 4.9%, greater than about 4.8%, greater than about 4.7%, greater than about 4.6%, greater than about 4.5%, greater than about 4.4%, greater than about 4.3%, greater than about 4.2%, greater than about 4.1%, greater than about 4.0%, greater than about 3.9%, greater than about 3.8%, greater than about 3.7%, or greater than about 3.6% of total risankizumab species with N-glycosylation. 【0079】 Embodiment 58. The composition of any one of embodiments 51 to 57, wherein the level of risankizumab containing high mannose N-glycans is about 3.6% to about 5.3%, about 3.6% to about 5.0%, about 3.6% to about 4.8%, about 3.6% to about 4.5%, about 3.6% to about 4.1%, about 3.6% to about 3.8%, about 3.8% to about 5.3%, about 4.1% to about 5.3%, about 4.5% to about 5.3%, about 4.8% to about 5.3%, about 5.0% to about 5.3%, about 4.3% to about 4.9%, or about 3.6% to about 4.9% of total risankizumab species with N-glycosylation. 【0080】 Embodiment 59. The composition of any one of embodiments 51 to 58, wherein the level of risankizumab containing the high mannose N-glycans is about 5.3%, about 5.2%, about 5.1%, about 5.0%, about 4.9%, about 4.8%, about 4.7%, about 4.6%, about 4.5%, about 4.4%, about 4.3%, about 4.2%, about 4.1%, about 4.0%, about 3.9%, about 3.8%, about 3.7%, or about 3.6% of total risankizumab species with N-glycosylation. 【0081】 Embodiment 60. The composition of embodiment 54, wherein the high mannose N-glycan is M5. 【0082】 Embodiment 61. The composition of embodiment 60, wherein the level of risankizumab comprising M5 is less than 5.3%, less than about 5.2%, less than about 5.1%, less than about 5.0%, less than about 4.9%, less than about 4.8%, less than about 4.7%, less than about 4.6%, less than about 4.5%, less than about 4.4%, less than about 4.3%, less than about 4.2%, less than about 4.1%, less than about 4.0%, less than 3.9%, less than about 3.8%, less than about 3.7%, less than about 3.6%, less than about 3.5%, less than about 3.4%, less than about 3.3%, less than about 3.2%, less than about 3.1%, less than about 3.0%, less than about 2.9%, or less than about 2.8% of total risankizumab species with N-glycosylation. 【0083】 Embodiment 62. The composition of embodiment 60 or 61, wherein the level of risankizumab comprising M5 is greater than about 5.2%, greater than about 5.1%, greater than about 5.0%, greater than about 4.9%, greater than about 4.8%, greater than about 4.7%, greater than about 4.6%, greater than about 4.5%, greater than about 4.4%, greater than about 4.3%, greater than about 4.2%, greater than about 4.1%, greater than about 4.0%, greater than about 3.9%, greater than about 3.8%, greater than about 3.7%, greater than about 3.6%, greater than about 3.5%, greater than about 3.4%, greater than about 3.3%, greater than about 3.2%, greater than about 3.1%, greater than about 3.0%, greater than about 2.9%, or greater than about 2.8%, or greater than about 2.7% of total risankizumab species with N-glycosylation. 【0084】 Embodiment 63. The composition of any one of embodiments 60-62, wherein the level of risankizumab containing M5 is between about 2.7% and about 5.2%, between about 3.1% and about 5.2%, between about 3.5% and about 5.2%, between about 4.0% and about 5.2%, between about 4.5% and about 5.2%, between about 5% and about 5.2%, between about 2.7% and about 5.0%, between about 2.7% and about 4.5%, between about 2.7% and about 4.0%, between about 2.7% and about 3.5%, between about 2.7% and about 3.1%, between about 3.2% and about 3.7%, or between about 2.7% and about 3.7% of total risankizumab species with N-glycosylation. 【0085】 Embodiment 64. The composition of any one of embodiments 60-63, wherein the level of risankizumab comprising M5 is about 2.7%, about 2.8%, about 2.9%, about 3.0%, about 3.1%, about 3.2%, about 3.3%, about 3.4%, about 3.5%, about 3.6%, about 3.7%, about 3.8%, about 3.9%, about 4.0%, about 4.1%, about 4.2%, about 4.3%, about 4.4%, about 4.5%, about 4.6%, about 4.7%, about 4.8%, about 4.9%, about 5.0%, about 5.1%, or about 5.2% of total risankizumab species with N-glycosylation. 【0086】 Embodiment 65. The composition of embodiment 54, wherein the high mannose glycan is M6. 【0087】 Embodiment 66. The composition of embodiment 65, wherein the level of risankizumab comprising M6 is less than about 2.6%, less than about 2.5%, less than about 2.4%, less than about 2.3%, less than about 2.2%, less than about 2.1%, less than about 2.0%, less than about 1.9%, less than about 1.8%, less than about 1.7%, less than about 1.6%, less than about 1.5%, less than about 1.4%, less than about 1.3%, less than about 1.2%, less than about 1.1%, less than about 1.0%, less than about 0.9%, less than about 0.8%, less than about 0.7%, less than about 0.6%, or less than about 0.5% of total risankizumab species with N-glycosylation. 【0088】 Embodiment 67. The composition of embodiment 65 or 66, wherein the level of risankizumab comprising M6 is greater than about 2.5%, greater than about 2.4%, greater than about 2.3%, greater than about 2.2%, greater than about 2.1%, greater than about 2.0%, greater than about 1.9%, greater than about 1.8%, greater than about 1.7%, greater than about 1.6%, greater than about 1.5%, greater than about 1.4%, greater than about 1.3%, greater than about 1.2%, greater than about 1.1%, greater than about 1.0%, greater than about 0.9%, greater than about 0.8%, greater than about 0.7%, greater than about 0.6%, greater than about 0.5%, or greater than about 0.4% of total risankizumab species with N-glycosylation. 【0089】 Embodiment 68. The composition of any one of embodiments 65-67, wherein the level of risankizumab containing M6 is between about 0.4% and about 2.5%, between about 0.4% and about 2.4%, between about 0.4% and about 2.2%, between about 0.4% and about 2.0%, between about 0.4% and about 1.8%, between about 0.4% and about 1.6%, between about 0.4% and about 1.4%, between about 0.4% and about 1.2%, between about 0.4% and about 1.0%, between about 0.4% and about 0.9%, between about 0.4% and about 0.8%, between about 0.4% and about 0.7%, between about 0.4% and about 0.6%, between about 0.4% and about 0.5%, or between about 0.6% and about 0.7% of total risankizumab species with N-glycosylation. 【0090】 Embodiment 69. The composition of any one of embodiments 65-68, wherein the level of risankizumab comprising M6 is about 2.5%, about 2.4%, about 2.3%, about 2.2%, about 2.1%, about 2.0%, about 1.9%, about 1.8%, about 1.7%, about 1.6%, about 1.5%, about 1.4%, about 1.3%, about 1.2%, about 1.1%, about 1.0%, about 0.9%, about 0.8%, about 0.7%, about 0.6%, about 0.5%, or about 0.4% of total risankizumab species with N-glycosylation. 【0091】 Embodiment 70. The composition of embodiment 54, wherein the high mannose glycan is M7. 【0092】 Embodiment 71. The composition of embodiment 70, wherein the level of risankizumab comprising M7 is less than about 2.0%, less than about 1.9%, less than about 1.8%, less than about 1.7%, less than about 1.6%, less than about 1.5%, less than about 1.4%, less than about 1.3%, less than about 1.2%, less than about 1.1%, less than about 1.0%, less than about 0.9%, less than about 0.8%, less than about 0.7%, less than about 0.6%, less than about 0.5%, or more than about 0.4% of total risankizumab species with N-glycosylation. 【0093】 Embodiment 72. The composition of embodiment 70 or 71, wherein the level of risankizumab comprising M7 is greater than about 1.9%, greater than about 1.8%, greater than about 1.7%, greater than about 1.6%, greater than about 1.5%, greater than about 1.4%, greater than about 1.3%, greater than about 1.2%, greater than about 1.1%, greater than about 1.0%, greater than about 0.9%, greater than about 0.8%, greater than about 0.7%, greater than about 0.6%, greater than about 0.5%, or greater than about 0.4% of total risankizumab species with N-glycosylation. 【0094】 Embodiment 73. The composition of any one of embodiments 70-72, wherein the level of risankizumab comprising M7 is from about 0.4% to about 1.9%, from about 0.4% to about 1.8%, from about 0.4% to about 1.6%, from about 0.4% to about 1.4%, from about 0.4% to about 1.2%, from about 0.4% to about 1.0%, from about 0.4% to about 0.9%, from about 0.4% to about 0.8%, from about 0.4% to about 0.7%, from about 0.4% to about 0.6%, from about 0.4% to about 0.5%, or from about 0.5% to about 0.6% of total risankizumab species with N-glycosylation. 【0095】 Embodiment 74. The composition of any one of embodiments 70-73, wherein the level of risankizumab comprising M7 is about 1.9%, about 1.8%, about 1.7%, about 1.6%, about 1.5%, about 1.4%, about 1.3%, about 1.2%, about 1.1%, about 1.0%, about 0.9%, about 0.8%, about 0.7%, about 0.6%, about 0.5%, or about 0.4% of total risankizumab species with N-glycosylation. 【0096】 Embodiment 75. The composition of any one of embodiments 51 to 74, wherein the level of risankizumab containing high mannose N-glycans is determined by 2-AB and HILIC-FL chromatography. 【0097】 Embodiment 76. The composition of any one of embodiments 51 to 74, wherein the level of risankizumab containing high mannose N-glycans is determined by RapiFluor HILIC-FL chromatography. 【0098】 Embodiment 77. The composition of any one of embodiments 51 to 76, wherein greater than about 84.4% of all risankizumab species with N-glycosylation have complex fucosylated oligosaccharides. 【0099】 Embodiment 78. The composition of embodiment 77, wherein about 88.0% to about 90.9% of all risankizumab species with N-glycosylation have fucosylated complex oligosaccharides. 【0100】 Embodiment 79. The composition of embodiment 77 or 78, wherein the level of risankizumab containing complex fucosylated oligosaccharides is determined by 2-AB and HILIC-FL chromatography. 【0101】 Embodiment 80. The composition of embodiment 77 or 78, wherein the level of risankizumab containing high mannose N-glycans is determined by RapiFluor HILIC-FL chromatography. 【0102】 Embodiment 81. The composition of any one of embodiments 51 to 80, wherein the composition comprises about 0.8% to about 1.4% aglycosylated risankizumab. 【0103】 Embodiment 82. The composition of embodiment 81, wherein the aglycosylated risankizumab is determined by tryptic peptide mapping. 【0104】 Embodiment 83. The composition of embodiment 51 or 52, having at least characteristic (b): at least about 99.1% of the risankizumab is present as monomers, and / or about 0.4% or less of the risankizumab is present as high molecular weight (HMW) species, as measured by ultra-performance size exclusion chromatography (UP-SEC). 【0105】 Embodiment 84. The composition of embodiment 83, wherein at least about 99.1% of the risankizumab is present as monomers and about 0.4% or less of the risankizumab is present as high molecular weight (HMW) species, as measured by ultra-performance size exclusion chromatography (UP-SEC). 【0106】 Embodiment 85. The composition of embodiment 83 or 84, wherein at least about 99.2%, at least about 99.3%, at least about 99.4%, at least about 99.5%, at least about 99.6%, or at least about 99.7% of the risankizumab is present as a monomer. 【0107】 Embodiment 86. The composition of any one of embodiments 83 to 85, wherein about 99.1% to about 99.7%, 99.1% to about 99.6%, about 99.2% to about 99.7%, or about 99.2% to about 99.6% of the risankizumab is present as a monomer. 【0108】 Embodiment 87. The composition of any one of embodiments 83-86, wherein no more than about 0.35%, no more than about 0.3%, no more than about 0.25%, no more than about 0.2%, no more than about 0.15%, or no more than about 0.1% of the risankizumab is present as high molecular weight (HMW) species. 【0109】 Embodiment 88. The composition of any one of embodiments 83 to 87, wherein about 0.1% to about 0.4%, about 0.1 to about 0.3%, about 0.1 to about 0.2%, about 0.2% to about 0.4%, or about 0.2% to about 0.3% of the risankizumab is present as a high molecular weight (HMW) species. 【0110】 Embodiment 89. The composition of embodiment 51 or 52, wherein the composition has at least characteristic (c): greater than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as low molecular weight (LMW) species, when measured by capillary gel electrophoresis under non-reducing conditions. 【0111】 Embodiment 90. The composition of embodiment 89, wherein greater than about 97.5% of the risankizumab is present as the main peak and less than about 2.2% of the risankizumab is present as low molecular weight (LMW) species. 【0112】 Embodiment 91. The composition of embodiment 89 or 90, wherein greater than about 97.6%, greater than about 97.7%, greater than about 97.8%, greater than about 97.9%, greater than about 98.0%, greater than about 98.1%, greater than about 98.2%, greater than about 98.3%, or greater than about 98.4% of the risankizumab is present as the main peak. 【0113】 Embodiment 92. The composition of any one of embodiments 89-91, wherein about 97.6% to about 98.4%, about 97.6% to about 98.3%, about 97.6% to about 98.2%, about 97.7% to about 98.4%, about 97.7% to about 98.3%, about 97.7% to about 98.2%, about 97.8% to about 98.4%, about 97.8% to about 98.3%, or about 97.8% to about 98.2% risankizumab is present as the main peak. 【0114】 Embodiment 93. The composition of any one of embodiments 89-92, wherein less than about 2.1%, less than 2.0%, less than 1.9%, less than 1.8%, less than 1.7%, less than 1.6%, or less than 1.5% of the risankizumab is present as low molecular weight (LMW) species. 【0115】 Embodiment 94. The composition of any one of embodiments 89-93, wherein about 1.5% to about 2.1%, about 1.6% to about 2.1%, about 1.7% to about 2.1%, about 1.5% to about 2.0%, about 1.6% to about 2.0%, or about 1.7% to about 2.0% of the risankizumab is present as a low molecular weight (LMW) species. 【0116】 Embodiment 95. The composition of embodiment 51 or 52, wherein the composition has at least characteristic (d), in which the incidence of treatment-emergent anti-drug antibodies (ADA) in a human is less than about 4.7% after administering a single 150 mg subcutaneous dose of the composition to the human. 【0117】 Embodiment 96. The composition of embodiment 95, wherein the incidence of treatment-emergent ADA is less than about 4.5%, less than about 4.0%, less than about 3.5%, less than about 3.0%, less than about 2.5%, less than about 2.0%, less than about 1.5%, less than about 1.0%, less than about 0.5%, less than about 0.4%, less than about 0.3%, less than about 0.2%, less than about 0.1%, less than about 0.01%, less than about 0.001%, or less than about 0.0001%. 【0118】 Embodiment 97. The composition of embodiment 95 or 96, wherein the incidence of treatment-emergent ADA is 0.0%. 【0119】 Embodiment 98. The composition of any one of embodiments 95 to 97, wherein the incidence of treatment-emergent ADA is measured after administering a single 150 mg subcutaneous dose of the composition to humans. 【0120】 Embodiment 99. The composition of any one of embodiments 95 to 98, wherein the presence of ADA is determined using a bridging electrochemiluminescence immunoassay. 【0121】 Embodiment 100. The composition of any one of embodiments 51 to 99, wherein the risankizumab is produced in a CHO cell line. 【0122】 Embodiment 101. The composition of any one of embodiments 51 to 100, further comprising a pharmaceutically acceptable excipient. 【0123】 Embodiment 102. The pharmaceutical composition of embodiment 101, wherein the excipient is selected from the group consisting of a surfactant, a polyol, or a buffer. 【0124】 Embodiment 103. The pharmaceutical composition of embodiment 102, wherein the polyol is trehalose. 【0125】 Embodiment 104. The pharmaceutical composition of embodiment 103, wherein the trehalose is present in an amount of about 150 to about 220 mM. 【0126】 Embodiment 105. The pharmaceutical composition of embodiment 104, wherein the trehalose is present in an amount of about 185 mM. 【0127】 Embodiment 106. The pharmaceutical composition of any one of embodiments 101 to 105, wherein the buffer is selected from the group consisting of acetate buffer and succinate buffer. 【0128】 Embodiment 107. The pharmaceutical composition of embodiment 106, wherein the buffer is an acetate buffer. 【0129】 Embodiment 108. The pharmaceutical composition of embodiment 107, wherein the acetate buffer is present in an amount of about 5 to about 50 mM. 【0130】 Embodiment 109. The pharmaceutical composition of embodiment 108, wherein the acetate buffer is present in an amount of about 10 mM. 【0131】 Embodiment 110. The pharmaceutical composition of any one of embodiments 101 to 109, wherein the surfactant is polysorbate 20 (PS20). 【0132】 Embodiment 111. The pharmaceutical composition of embodiment 110, wherein the PS20 is present in an amount of about 0.02 mg / ml. 【0133】 Embodiment 112. The pharmaceutical composition of embodiment 111, wherein the PS20 is present in an amount of about 0.2 mg / mL. 【0134】 Embodiment 113. The pharmaceutical composition of embodiment 112, comprising 150 mg / ml risankizumab; 185 mM trehalose; 10 mM acetic acid; and 0.20 mg / mL polysorbate 20, wherein the pH is about 5.7. 【0135】 Embodiment 114. The pharmaceutical composition of embodiment 112, comprising 150 mg / ml risankizumab; 0.054 mg / mL acetic acid; 1.24 mg / mL sodium acetate trihydrate; 70 mg / mL trehalose dihydrate; 0.20 mg / mL polysorbate 20; and water for injection, USP, having a pH of about 5.7. 【0136】 Embodiment 115. The pharmaceutical composition of embodiment 110, wherein the risankizumab is present in a concentration of 150 mg / ml. 【0137】 Embodiment 116. The pharmaceutical composition of any one of embodiments 101 to 115, wherein the pharmaceutical composition is packaged in a vial, a pre-filled syringe, or an on-body device. 【0138】 Embodiment 117. The pharmaceutical composition of any one of embodiments 101 to 116, wherein the pharmaceutical composition is suitable for subcutaneous injection. 【0139】 Embodiment 118. The pharmaceutical composition of any one of embodiments 101 to 117, wherein the pharmaceutical composition is suitable for intravenous injection. 【0140】 Embodiment 119. The pharmaceutical composition of any one of embodiments 101 to 118, wherein the pharmaceutical composition is a liquid composition. 【0141】 Embodiment 120. The pharmaceutical composition of any one of embodiments 101 to 118, wherein the pharmaceutical composition is an aqueous liquid composition. 【0142】 Embodiment 121. A method of treating an immunological disease with the pharmaceutical composition of any one of embodiments 101 to 120. 【0143】 Embodiment 122. A process for producing a risankizumab drug product having an amount of PLA2 that is less than about 250 pg per mg of risankizumab, comprising: (1) culturing a host cell line that expresses risankizumab in a growth medium under conditions that allow production of risankizumab; (2) clarifying the growth medium by centrifugation and depth filtration; (3) contacting the clarified medium containing risankizumab with a Protein A resin; (4) eluting risankizumab from the Protein A resin to obtain a first eluate; and (5) contacting the first eluate with a Protein A resin. (6) contacting the filtered first eluate with a mixed-mode resin to obtain a first flow-through containing risankizumab; (7) contacting the first flow-through with a cation exchange resin; (8) eluting risankizumab from the cation exchange resin to obtain a second eluate; and (9) treating the second eluate by ultrafiltration and dialysis, thereby obtaining a risankizumab drug product having an amount of PLA2 that is less than about 250 pg per mg of risankizumab. 【0144】 Embodiment 123. The process of embodiment 122, further comprising subjecting the first eluate to a viral inactivation step prior to step (5). 【0145】 Embodiment 124. The process of embodiment 122 or 123, further comprising subjecting the second eluate to viral filtration prior to step (9). 【0146】 Embodiment 125. A process for producing a risankizumab drug product having risankizumab containing high-mannose N-glycans in an amount that is less than about 5.4% of total risankizumab species with N-glycosylation, comprising: (1) culturing a host cell line that expresses risankizumab in a growth medium under conditions that allow for the production of risankizumab; (2) clarifying the growth medium by centrifugation and depth filtration; (3) contacting the clarified medium containing risankizumab with a Protein A resin; (4) eluting risankizumab from the Protein A resin to obtain a first eluate; and (5) obtaining a first eluate. (6) contacting the filtered first eluate with a mixed-mode resin to obtain a first flow-through containing risankizumab; (7) contacting the first flow-through with a cation exchange resin; (8) eluting risankizumab from the cation exchange resin to obtain a second eluate; and (9) treating the second eluate by ultrafiltration and dialysis, thereby obtaining a risankizumab drug product having risankizumab containing high-mannose N-glycans in an amount that is less than about 5.4% of total risankizumab species with N-glycosylation. 【0147】 Embodiment 126. The process of embodiment 125, further comprising subjecting the first eluate to a viral inactivation step prior to step (5). 【0148】 Embodiment 127. The process of embodiment 125 or 126, further comprising subjecting the second eluate to viral filtration prior to step (9). 【0149】 Embodiment 128. A composition comprising (1) risankizumab and (2) poloxamer 188 (P188), optionally without polysorbate 20 (PS20) and / or polysorbate 80 (PS80). 【0150】 Embodiment 129. The composition of embodiment 128, comprising about 60 mg / ml to about 150 mg / ml of risankizumab. 【0151】 Embodiment 130. The composition of embodiment 128 or 129, further comprising phospholipase A2 (PLA2). 【0152】 Embodiment 131. The composition of any one of embodiments 128 to 130, wherein the PLA2 is PLA2G15. 【0153】 Embodiment 132. The composition of any one of embodiments 128-131, wherein the level of PLA2 is greater than about 250 pg / mg of risankizumab, and the level of PLA2 is greater than about 260 pg, greater than about 270 pg, greater than about 280 pg, greater than about 290 pg, greater than about 300 pg, greater than about 310 pg, greater than about 320 pg, greater than about 330 pg, greater than about 340 pg, greater than about 350 pg, greater than about 360 pg, greater than about 380 pg, greater than about 400 pg, greater than about 450 pg, greater than about 500 pg, greater than about 550 pg, greater than about 600 pg, greater than about 650 pg, greater than about 700 pg, greater than about 750 pg, greater than about 800 pg, greater than about 900 pg, or greater than about 1000 pg. 【0154】 Embodiment 133. The level of PLA2 is from about 250 pg to about 1100 pg, from about 260 pg to about 1100 pg, from about 270 pg to about 1100 pg, from about 280 pg to about 1100 pg, from about 290 pg to about 1100 pg, from about 300 pg to about 1100 pg, from about 310 pg to about 1100 pg, from about 320 pg to about 1100 pg, from about 340 pg to about 1100 pg, from about 360 pg to about 1100 pg, or from about 250 pg to about 1100 pg per mg of risankizumab. 132. The composition of any one of embodiments 128-131, wherein the ATP concentration is 0 pg to about 1000 pg, about 250 pg to about 900 pg, about 250 pg to about 800 pg, about 250 pg to about 700 pg, about 250 pg to about 600 pg, about 250 pg to about 500 pg, about 250 pg to about 400 pg, about 250 pg to about 1030 pg, about 290 pg to about 1090 pg, about 360 pg to about 450 pg, or about 310 pg to about 920 pg. 【0155】 Embodiment 134. The composition of any one of embodiments 128 to 131, wherein the level of PLA2 is about 260 pg, about 270 pg, about 280 pg, about 290 pg, about 300 pg, about 310 pg, about 320 pg, about 330 pg, about 340 pg, about 350 pg, about 360 pg, about 380 pg, about 400 pg, about 450 pg, about 500 pg, about 550 pg, about 600 pg, about 650 pg, about 700 pg, about 750 pg, about 800 pg, about 900 pg, about 1000 pg, or about 1100 pg per mg of risankizumab. 【0156】 Embodiment 135. The composition of any one of embodiments 130 to 134, wherein the level of PLA2 is determined by ELISA. 【0157】 Embodiment 136. The composition of any one of embodiments 128-135, wherein at least 85% of the initial amount of P118 is retained after 6 months of storage at 5°C. 【0158】 Embodiment 13 The composition of any one of embodiments 128-135, wherein at least 80% of the initial amount of P118 is retained after 6 months of storage at 7.5°C. 【0159】 Embodiment 138. The composition of any one of embodiments 128-135, wherein at least 65% of the initial amount of P118 is retained after storage at 25°C for 3 months. 【0160】 Embodiment 139. The composition of any one of embodiments 128-135, wherein at least 60% of the initial amount of P118 is retained after 6 months of storage at 25°C. 【0161】 Embodiment 140. The composition of any one of embodiments 128-135, wherein at least 60% of the initial amount of P118 is retained after storage at 40°C for 3 months. 【0162】 Embodiment 141. The composition of any one of embodiments 128-135, wherein at least 60% of the initial amount of P118 is retained after storage at 40°C for 6 months. 【0163】 Embodiment 142. The composition of any one of embodiments 136 to 141, wherein the P188 is measured using a Pluronic F-68 colorimetric assay. 【0164】 Further advantages of the present disclosure will be apparent to those skilled in the art upon reading this patent application. The embodiments of the present disclosure described in the following paragraphs are intended to illustrate the present invention and should not be considered to narrow the scope of the present invention. 【0165】 definition The singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. 【0166】 The term "and / or" as used herein in phrases such as "A and / or B" is intended to mean "A and B," "A or B," "A" or "B." 【0167】 The term "about" generally refers to a range of numbers that one of ordinary skill in the art would consider equivalent to the recited value (i.e., having the same function or result). In many instances, the term "about" may include numbers rounded to the nearest significant figure. 【0168】 Unless the context otherwise requires, the terms "comprise", "comprises", and "comprising" are used with the clear understanding that they are to be interpreted inclusively and not exclusively, to indicate the inclusion of recited features but not the exclusion of one or more other such features. 【0169】 The term "carrier" used in conjunction with pharmaceutical excipients refers to any and all solvents, dispersion media, preservatives, coatings, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration. 【0170】 The terms "patient," "subject," "individual," and the like refer to a human. 【0171】 Risankizumab According to USAN, risankizumab has the following chemical name: 【0172】 1. Immunoglobulin G1, anti-(human interleukin-23 subunit p19) (human-Mus musculus heavy chain), disulfide dimer with human-Mus musculus kappa chain 【0173】 2. Immunoglobulin G1-kappa, anti-(human interleukin-23 subunit alpha (IL-23-A, interleukin-23 subunit p19, IL-23p19)), humanized monoclonal antibody, gamma 1 heavy chain (1-449) [humanized VH (Homo sapiens IGHV1-69*08 (79%)-(IGHD)-IGHJ6*01 (91%)) [8.8.13] (1-120)-Homo sapiens IGHG1*03{CH2 L 4 >A(237), L 5 >A(238), CH3 K 107>-(450)}(121-449)], (223-214')-disulfide and kappa light chain (1'-214') [humanized V-KAPPA (Homo sapiens IGKV1-27*01 (80%)-IGKJ2*02 (91%)) [6.3.9] (1'-107')-Homo sapiens IGKC*01 (108'-214')]; dimer (229-229":232-232")-bisdisulfide 【0174】 According to INN (see WHO Drug Information, 29 (2), 254-255), risankizumab has the following chemical name: 【0175】 Immunoglobulin G1-kappa, anti-[Homo sapiens IL23A (interleukin-23 subunit alpha, IL-23A, IL23 subunit p19 IL23p19)], humanized monoclonal antibody, gamma 1 heavy chain (1-449) [humanized VH (Homo sapiens IGHV1-69*02 (79.40%)-(IGHD)-IGHJ5*01) [8.8.13] (1-120)-Homo sapiens IGHG1*01, G1m17,1 (CH1 (121-218), hinge (219-233), CH2 L1.3>A (237), L1.2>A (238) (234-343), CH3 (344-448), CHS K2>del(449)(121-449)], (223-214')-disulfide and kappa light chain (1'-214') [humanized V-KAPPA (Homo sapiens IGKV1-27*01(80.00%)-IGKJ2*01)[6.3.9](1'-107')-Homo sapiens IGKC*01, Km3(108'-214')], dimer (229-229":232-232")-bisdisulfide 【0176】 Risankizumab binds human IL-23 with high affinity, with an inhibitory concentration (IC) of less than 10 pM compared to 167 pM for ustekinumab in the same system. 50Risankizumab inhibits IL-23-stimulated IL-17 production at the highest concentration tested (33 nM). Risankizumab has no effect on IL-12 and does not inhibit IL-12-stimulated IFN-γ production at the highest concentration tested (33 nM). 【0177】 Risankizumab amino acid sequence Risankizumab has the CDRs shown in Tables 1 and 2. The variable regions of risankizumab are shown in Table 3. 【0178】 [Table 1] 【0179】 [Table 2] 【0180】 [Table 3] 【0181】 Risankizumab contains the heavy and light chain sequences shown in Table 4. 【0182】 [Table 4] 【0183】 Risankizumab compositions with reduced hitchhiker proteins In one embodiment, the present disclosure relates to a liquid composition comprising: (1) risankizumab; and (2) PLA2 in an amount that is less than about 250 pg per mg of risankizumab. 【0184】 In one embodiment, the liquid composition described herein contains about 60 mg / ml to about 150 mg / ml of risankizumab. For example, but not limited to, the liquid composition described herein may contain about 70 mg / ml to about 150 mg / ml, about 80 mg / ml to about 150 mg / ml, about 90 mg / ml to about 150 mg / ml, about 100 mg / ml to about 150 mg / ml, about 110 mg / ml to about 150 mg / ml, about 120 mg / ml to about 150 mg / ml, about 130 mg / ml to about 150 mg / ml, or about 140 mg / ml to about 150 mg / ml. / ml, 60 mg / ml to about 70 mg / ml, 60 mg / ml to about 80 mg / ml, 60 mg / ml to about 90 mg / ml, 60 mg / ml to about 100 mg / ml, 60 mg / ml to about 110 mg / ml, 60 mg / ml to about 120 mg / ml, 60 mg / ml to about 130 mg / ml, or 60 mg / ml to about 140 mg / ml of risankizumab, and ranges and amounts between any of these aforementioned concentrations. 【0185】 In one embodiment, the liquid compositions described herein contain about 60 mg / ml, about 70 mg / ml, about 80 mg / ml, about 90 mg / ml, about 100 mg / ml, about 110 mg / ml, about 120 mg / ml, about 130 mg / ml, about 140 mg / ml, or about 150 mg / ml of risankizumab. 【0186】 As used herein, the term "phospholipase A2" or "PLA2" refers to a well-known family of enzymes that catalyze the hydrolysis of membrane phospholipids. PLA2 catalyzes the hydrolysis of membrane glycerophospholipids at the sn-2 position to liberate arachidonic acid (AA), a precursor of eicosanoids, including prostaglandins (PGs) and leukotrienes (LTs). The same reaction also produces lysophospholipids, which represent another class of lipid mediators (Murakami and Kudo (2002) J. Biochem 131:285-292). There are at least 16 groups of phospholipase A2s. Dennis and colleagues have classified them into six groups based on their properties. These include secreted phospholipase A2 (sPLA2 groups I, II, III, V, IX, X, XI, XII, XIII, and XIV), cytosolic phospholipase A2 (group IV cPLA2), calcium-independent phospholipase A2 (group VI iPLA2), PAF acetylhydrolase (GVII and GVIII pAF-AH pLA2), lysosomal phospholipase A2 (group XV LPLA2), and fat-specific phospholipase A2 (GXVI AdPLA) (Shayman and Tesmer (2019) Molecular and Cell Biology of Lipids 1864:932-940). In some embodiments, a PLA2 according to the present disclosure can catalyze the hydrolysis of surfactants such as polysorbate 20 (PS20), polysorbate 80 (PS80), polysorbate 40 (PS40), polysorbate 60 (PS60), polysorbate 65 (PS65), or poloxamer 188. Exemplary PLA2s according to the present disclosure include, but are not limited to, PLA2G15, PLA2G7, and PLA2G2. 【0187】 As used herein, the term "PLA2G15," also known as "Group XV PLA2," refers to a unique member of the PLA2 family (Shayman et al. (2011) prog. Lipid Res. 50:1-13). PLA2G15 is localized intracellularly to lysosomes and late endosomes, has an acidic pH optimum, and functions as a PLA2 (Abe and Shayman (2007) J. Lipid Res. 48:2255-2263). The primary structure of PLA2G15 is highly conserved between mouse, bovine, and human. The gene contains six exons. The primary structure of the human and mouse enzyme consists of 412 amino acids (compared to 407 for the bovine enzyme). This enzyme contains a consensus sequence including a signal peptide cleavage site and the lipase motif AXSXG, characteristic of serine hydrolases. The serine is part of the catalytic triad, and it also contains aspartic acid and histidine. An amino-terminal 33 amino acid signal peptide is present, with a cleavage site between proline 33 and alanine 34 in the mouse and human peptides. Additionally, four N-linked glycosylation sites are present in the mouse and human protein (three in the bovine protein) (Hiraoka and Shayman (2005) J. Lipid Res. 46:2441-2447). The structure and function of PLA2G15 are further described in Shayman and Tesmer, Molecular and Cell Biology of Lipids (2019) 1864:932-940, the contents of which are incorporated herein by reference in their entirety. 【0188】 Representative human PLA2G15 cDNA and human PLA2G15 protein sequences are known in the art and are published by the National Center for Biotechnology Information (NCBI). For example, two distinct human PLA2G15 isoforms are known. Human PLA2G15 isoform 1 (NP_036452.1) can be encoded by transcript variant 1 (NM_012320.4), a longer transcript. Human PLA2G15 isoform 2 (NP_001350480.1) can be encoded by transcript variant 2 (NM_001363551.2), which is shorter than isoform 1 and has a distinct C-terminus. Nucleic acid and polypeptide sequences of PLA2G15 orthologs from organisms other than humans are known, e.g., chimpanzee PLA2G15 (XM_001167383.5 and XP_001167383.1), rhesus monkey PLA2G15 (NM_001265818.1 and NP_001252747.1), bovine PLA2G15 (NM_174560.2 and NP_776985.2), canine PLA2G15 (NM_001002940.1 and NP_001002940.1), rat PLA2G15 (NM_001004277.2 and NP_001004277.1), mouse PLA2G15 (NM_001357319.1 and NP_00134424 8.1; NM_133792.3 and NP_598553.1), Chinese hamster PLA2G15 (XM_003504311.5 and XP_003504359.1; XM_027437910.2 and XP_027293711.1), chicken PLA2G15 (XM_001231518.7 and XP_0012 31519.1), Xenopus tropicalis PLA2G15 (XM_012962222.3 and XP_012817676.2; XM_031900713.1 and XP_031756573.1), and zebrafish PLA2G15 (NM_001386706.1 and NP_001373635.1). Representative sequences of PLA2G15 orthologs are presented in Table 5. 【0189】 Suitable anti-PLA2G15 antibodies for detecting PLA2G15 protein are known in the art, e.g., antibody catalog numbers NBP1-92089, H00023659-M01, and NBP2-17193, NBP1-92088, and NBP2-17192 (Novus Biologicals, Littleton, CO), antibody orb185108 (biorbyt, St. Louis, MO), antibody catalog numbers ABIN7004525, ABIN2580837, ABIN2580838, and ABIN2580836 (available on the World Wide Web at antibodies-online.com), antibody catalog number sc Examples of PLA2G15-specific antibodies include sc-376078, sc-529817, sc-543705, sc-522840, sc-376078AC, sc-376078HRP, sc-376078FITC, sc-376078PE, and sc-376078AF488 (Santa Cruz Biotechnology, Dallas, TX). Additionally, reagents for detecting PLA2G15 expression are also well known. Several clinical tests for PLA2G15 are available on the NIH Genetic Testing Registry (GTR®) (e.g., GTR Test ID: GTR000543805.3, provided by Fulgent Clinical Diagnostics Lab, Temple City, CA). 【0190】 [Table 5] 【0191】 The * in Table 5 refers to a polypeptide molecule containing an amino acid sequence, or a portion thereof, that has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5% or more identity over the entire length including the amino acid sequence of any of the SEQ ID NOs listed in Table 5. Such polypeptides can have the function of the full-length polypeptides described further herein. Further included are polypeptides with or without a signal peptide, and / or that include only the proprotein and / or that include only the mature protein. 【0192】 In one embodiment, the liquid compositions described herein contain a detectable amount of PLA2 that is less than 250 pg per mg of risankizumab. For example, and without limitation, the PLA2 in the liquid compositions described herein may be less than about 240, less than about 220, less than about 200, less than about 180, less than about 160, less than about 140, less than about 120, less than about 100, less than about 90, less than about 80, less than about 70, less than about 60, less than about 50, less than about 40, less than about 30, less than about 25, less than about 20, less than about 15, less than about 10, less than about 9, less than about 8, less than about 7, less than about 6, less than about 5, less than about 4.4, less than about 3, less than about 2, less than about 1, less than about 0.5, or less than about 0.1 pg per mg of risankizumab. less than about 0.05, or less than about 0.01 pg, or any range therebetween (e.g., including about 200 to about 249, about 160 to about 200, about 120 to about 160, about 100 to about 120, about 80 to about 100, about 60 to about 80, about 40 to about 60, about 25 to about 40, about 10 to about 25, about 5 to about 10, about 4 to about 10, about 1 to about 5, about 1 to about 4, about 1 to about 3, about 1 to about 2, about 0.5 to about 1, about 0.1 to about 0.5, about 0.05 to about 0.1, or about 0.01 to about 0.5 pg per mg of risankizumab). In some embodiments, PLA2 is present in an amount that is below or at the limit of detection of a PLA2 detection assay, e.g., less than about 9 pg / mg of risankizumab, or less than about 4.4 pg / mg of risankizumab. In one embodiment, PLA2 is present in an amount that is about 70 to about 240 pg / mg of risankizumab. 【0193】 In one embodiment, the liquid composition described herein comprises PLA2 in an amount that is about 240, about 220, about 200, about 180, about 160, about 140, about 120, about 100, about 90, about 80, about 70, about 60, about 50, about 40, about 30, about 25, about 20, about 15, about 10, about 9, about 8, about 7, about 6, about 5, about 4.4, about 3, about 2, about 1, about 0.5, about 0.1, about 0.05, or about 0.01 pg per mg of risankizumab. 【0194】 In one embodiment, the liquid compositions described herein contain less than about 250 pg of risankizumab per mg, but more than about 240 pg, more than about 220 pg, more than about 200 pg, more than about 180 pg, more than about 160 pg, more than about 140 pg, more than about 120 pg, more than about 100 pg, more than about 90 pg, more than about 80 pg, more than about 70 pg, more than about 60 pg, more than about 50 pg, more than about 60 pg, more than about 70 pg, more than about 80 pg, more than about 9 ... pg, greater than about 40 pg, greater than about 30 pg, greater than about 25 pg, greater than about 20 pg, greater than about 15 pg, greater than about 10 pg, greater than about 9 pg, greater than about 8 pg, greater than about 7 pg, greater than about 6 pg, greater than about 5 pg, greater than about 4 pg, greater than about 3 pg, greater than about 2 pg, greater than about 1 pg, greater than about 0.5 pg, greater than about 0.1 pg, greater than about 0.05 pg, or greater than about 0.01 pg. 【0195】 In one embodiment, the disclosure relates to a liquid composition comprising: (1) about 150 mg / ml of risankizumab, the liquid composition comprising a light chain having the amino acid sequence of SEQ ID NO: 9 and a heavy chain having the amino acid sequence of SEQ ID NO: 10; and (2) an amount of PLA2 that is less than about 250 pg (e.g., less than 240, about 70 to about 240, less than about 9, or less than about 4.4) per mg of risankizumab. 【0196】 The PLA2 in the liquid compositions described herein can be PLA2G2, PLA2G15, or a combination thereof. In one embodiment, the PLA2 is PLA2G15. 【0197】 The amount of PLA2 in the liquid compositions described herein can be determined using methods known in the art, for example, by mass spectrometry or by ELISA. In one embodiment, the amount of PLA2 in the liquid compositions described herein is determined by ELISA, for example, by using the ELISA method described in Example 9. 【0198】 In one embodiment, the PLA2 in the liquid compositions described herein is derived from a CHO cell line. 【0199】 In one embodiment, the liquid compositions described herein further comprise one or more of a surfactant, a polyol, and a buffer. 【0200】 The polyol may be selected from the group consisting of trehalose, mannitol, sucrose, and sorbitol. In one embodiment, the polyol is trehalose, and the trehalose is present in an amount of about 150 to about 220 mM (e.g., about 185 mM). 【0201】 The buffer may be selected from the group consisting of acetate buffer, histidine buffer, citrate buffer, phosphate buffer, glycine buffer, and arginine buffer. In one embodiment, the buffer is acetate buffer, and the acetate buffer is present in an amount of about 5 to about 100 mM (e.g., about 10 mM). 【0202】 The surfactant may be selected from the group consisting of polysorbate 20 (PS20), polysorbate 80 (PS80), polysorbate 40 (PS40), polysorbate 60 (PS60), polysorbate 65 (PS65), and poloxamer 188. In one embodiment, the surfactant is PS20, and the PS20 is present in an amount of up to 1.0 mg / ml (e.g., about 1.0 mg / ml, about 0.8 mg / ml, about 0.6 mg / ml, about 0.4 mg / ml, about 0.2 mg / ml, or about 0.1 mg / ml). In another embodiment, the surfactant is PS80, and the PS80 is present in an amount of up to 1.0 mg / ml (e.g., about 1.0 mg / ml, about 0.8 mg / ml, about 0.6 mg / ml, about 0.4 mg / ml, about 0.2 mg / ml, or about 0.1 mg / ml). 【0203】 In one embodiment, the liquid compositions described herein have a pH of about 5.0 to about 6.5 (eg, about 5.7). 【0204】 In one embodiment, the liquid composition described herein comprises: (1) 150 mg / mL risankizumab comprising a light chain having the amino acid sequence of SEQ ID NO:9 and a heavy chain having the amino acid sequence of SEQ ID NO:10, 185 mM trehalose, 10 mM acetate, and 0.20 mg / mL polysorbate 20, wherein the liquid composition has a pH of about 5.7; and (2) an amount of PLA2 (e.g., PLA2G15) that is less than about 250 pg per mg of risankizumab. 【0205】 In one embodiment, the liquid composition described herein comprises: (1) 150 mg / mL risankizumab comprising a light chain having the amino acid sequence of SEQ ID NO:9 and a heavy chain having the amino acid sequence of SEQ ID NO:10, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, wherein the liquid composition has a pH of about 5.7; and (2) an amount of PLA2 (e.g., PLA2G15) that is less than about 250 pg per mg of risankizumab. 【0206】 In one embodiment, the liquid composition described herein comprises: (1) 150 mg / mL risankizumab comprising a light chain having the amino acid sequence of SEQ ID NO:9 and a heavy chain having the amino acid sequence of SEQ ID NO:10, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, wherein the liquid composition has a pH of about 5.7; and (2) an amount of PLA2 (e.g., PLA2G15) that is less than about 250 pg per mg of risankizumab (e.g., less than or equal to 240, between about 70 and about 240, less than about 9, or less than about 4.4). 【0207】 In one embodiment, the liquid composition described herein comprises: (1) 150 mg / mL risankizumab comprising a light chain having the amino acid sequence of SEQ ID NO:9 and a heavy chain having the amino acid sequence of SEQ ID NO:10, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20 (the liquid composition has a pH of about 5.7); and (2) an amount of PLA2 (e.g., PLA2G15) that is less than about 250 pg (e.g., less than 240, between about 70 and about 240, less than about 9, or less than about 4.4) per mg of risankizumab (the amount of PLA2 is determined by ELISA (e.g., the ELISA method described in Example 9)). 【0208】 Liquid formulations encompassed by the present disclosure may contain added water, such as USP grade water. 【0209】 In some embodiments, the liquid pharmaceutical formulations described herein are packaged in a vial, a pre-filled syringe, or an on-body device. 【0210】 In some embodiments, the liquid pharmaceutical formulations described herein are suitable for parenteral administration.Parenteral administration includes, for example, subcutaneous, intramuscular, intradermal, intramedullary injection, as well as intrathecal, direct intracerebroventricular, intravenous, intraperitoneal, and intravitreal injection.In one embodiment, the disclosed liquid formulations are injectable formulations.In one embodiment, the disclosed liquid formulations are suitable for subcutaneous or intravenous injection. 【0211】 Risankizumab compositions with reduced high mannose N-glycans, increased purity, and / or reduced immunogenicity In another aspect, the disclosure relates to a composition comprising risankizumab, wherein the composition has one or more of the following characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans; (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as high molecular weight (HMW) species, as measured by ultra-performance size-exclusion chromatography (UP-SEC); (c) more than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as low molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR); and / or (d) the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% following administration of a single 150 mg subcutaneous dose of the composition to humans. 【0212】 In one embodiment, the composition is a pharmaceutical composition. 【0213】 In one embodiment, the pharmaceutical composition is a liquid composition. 【0214】 In one embodiment, the pharmaceutical composition is an aqueous composition. 【0215】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has characteristic (a): less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans. 【0216】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has characteristic (b): at least about 99.1% of the risankizumab is present as monomers, and / or about 0.4% or less of the risankizumab is present as high molecular weight (HMW) species, as measured by ultra-performance size exclusion chromatography (UP-SEC). 【0217】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has characteristic (c): greater than about 97.5% of the risankizumab is present as a main peak, and / or less than about 2.2% of the risankizumab is present as low molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR). 【0218】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has characteristic (d): an incidence of treatment-emergent anti-drug antibodies (ADA) of less than about 4.7% following administration of a single 150 mg subcutaneous dose of the composition to a human. 【0219】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans; and (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by ultra-performance size-exclusion chromatography (UP-SEC). 【0220】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the pharmaceutical composition has the characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans; and (c) greater than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as low molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR). 【0221】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans; and (d) the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% after administration of a single 150 mg subcutaneous dose of the composition. 【0222】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics: (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by ultra-performance size-exclusion chromatography (UP-SEC); and (c) greater than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as a low molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR). 【0223】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics: (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by ultra-performance size exclusion chromatography (UP-SEC); and (d) the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% after a single subcutaneous dose of the composition. 【0224】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics (c) greater than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as a small molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR), and (d) the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% after a single subcutaneous dose of the composition. 【0225】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans; (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by ultra-performance size-exclusion chromatography (UP-SEC); and (c) more than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as a low molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR). 【0226】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans; (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by ultra-performance size-exclusion chromatography (UP-SEC); and (d) the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% following administration of a single 150 mg subcutaneous dose of the composition to humans. 【0227】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics: (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by ultra-performance size-exclusion chromatography (UP-SEC); (c) greater than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as a low molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR); and (d) the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% following subcutaneous administration of a single dose of the composition to humans. 【0228】 In one embodiment, the disclosure relates to a composition comprising risankizumab, wherein the composition has the characteristics: (a) less than about 5.4% of all risankizumab species with N-glycosylation have high-mannose N-glycans; (b) at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as high molecular weight (HMW) species, as measured by ultra-performance size-exclusion chromatography (UP-SEC); (c) more than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as low molecular weight (LMW) species, as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR); and (d) the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% following subcutaneous administration of a single dose of the composition to humans. 【0229】 In one embodiment, the compositions described herein contain about 60 mg / ml to about 150 mg / ml of risankizumab. For example, but not limited to, the compositions described herein may contain about 70 mg / ml to about 150 mg / ml, about 80 mg / ml to about 150 mg / ml, about 90 mg / ml to about 150 mg / ml, about 100 mg / ml to about 150 mg / ml, about 110 mg / ml to about 150 mg / ml, about 120 mg / ml to about 150 mg / ml, about 130 mg / ml to about 150 mg / ml, or about 140 mg / ml to about 150 mg / ml. , 60 mg / ml to about 70 mg / ml, 60 mg / ml to about 80 mg / ml, 60 mg / ml to about 90 mg / ml, 60 mg / ml to about 100 mg / ml, 60 mg / ml to about 110 mg / ml, 60 mg / ml to about 120 mg / ml, 60 mg / ml to about 130 mg / ml, or 60 mg / ml to about 140 mg / ml of risankizumab, and ranges and amounts between any of these aforementioned concentrations. 【0230】 In one embodiment, the compositions described herein contain about 60 mg / ml, about 70 mg / ml, about 80 mg / ml, about 90 mg / ml, about 100 mg / ml, about 110 mg / ml, about 120 mg / ml, about 130 mg / ml, about 140 mg / ml, or about 150 mg / ml of risankizumab. 【0231】 In one embodiment, risankizumab is produced in a CHO cell line. 【0232】 In one embodiment, the compositions described herein further comprise a pharmaceutically acceptable excipient. 【0233】 In one embodiment, a pharmaceutical composition described herein comprises 150 mg / ml risankizumab, 185 mM trehalose, 10 mM acetic acid, and 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7. 【0234】 In one embodiment, a pharmaceutical composition described herein comprises 150 mg / ml risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, water for injection, USP, wherein the pharmaceutical composition has a pH of about 5.7. 【0235】 In one embodiment, the pharmaceutical compositions described herein have a pH of about 5.0 to about 6.5 (eg, about 5.7). 【0236】 In some embodiments, the pharmaceutical compositions described herein are packaged in a vial, a pre-filled syringe, or an on-body device. 【0237】 In some embodiments, the pharmaceutical compositions described herein are suitable for parenteral administration.Parenteral administration includes, for example, subcutaneous injection, intramuscular injection, intradermal injection, intramedullary injection, intrathecal injection, direct intraventricular injection, intravenous injection, intraperitoneal injection, and intravitreal injection.In one embodiment, the disclosed liquid formulation is an injectable formulation.In one embodiment, the disclosed liquid formulation is suitable for subcutaneous or intravenous injection. 【0238】 In some aspects, provided herein are methods of treating immunological diseases using the compositions described herein, including, but not limited to, autoimmune and inflammatory diseases (e.g., psoriasis, inflammatory bowel disease, ulcerative colitis, psoriatic arthritis, and Crohn's disease). 【0239】 a. Risankizumab pharmaceutical compositions containing reduced high-mannose N-glycans In one embodiment, the compositions provided herein have at least the characteristic (a) that less than about 5.4% of all risankizumab species with N-glycosylation have high mannose N-glycans. 【0240】 In one embodiment, the high mannose N-glycan comprises one or more high mannose N-glycans selected from mannose 5 N-glycan (M5), mannose 6 N-glycan (M6), and mannose 7 N-glycan (M7). For example, in one embodiment, the high mannose N-glycans are M5, M6, and M7. 【0241】 In one embodiment, the compositions described herein comprise multiple risankizumab species with or without N-glycosylation, wherein the sum of M5-containing risankizumab, M6-containing risankizumab, and M7-containing risankizumab is less than 5.4% of the total N-glycosylated risankizumab species. For example, and without limitation, the sum of risankizumab with M5, risankizumab with M6, and risankizumab with M7 in the compositions described herein is less than about 5.3%, less than about 5.2%, less than about 5.1%, less than about 5.0%, less than about 4.9%, less than about 4.8%, less than about 4.7%, less than about 4.6%, less than about 4.5%, less than about 4.4%, less than about 4.3%, less than about 4.2%, less than about 4.1%, less than about 4.0%, less than about 3.9%, less than about 3.8%, or less than about 3. 0.7%, or any range therebetween (inclusive), e.g., about 3.6% to about 5.3%, about 3.6% to about 5.0%, about 3.6% to about 4.8%, about 3.6% to about 4.5%, about 3.6% to about 4.1%, about 3.6% to about 3.8%, about 3.8% to about 5.3%, about 4.1% to about 5.3%, about 4.5% to about 5.3%, about 4.8% to about 5.3%, about 5.0% to about 5.3%, about 4.3% to about 4.9%, or about 3.6% to about 4.9% of total risankizumab species with N-glycosylation. 【0242】 In one embodiment, the compositions described herein comprise the sum of risankizumab with M5, risankizumab with M6, and risankizumab with M7 in an amount that is less than about 5.4% of total risankizumab species with N-glycosylation, but greater than about 5.3%, greater than about 5.2%, greater than about 5.1%, greater than about 5.0%, greater than about 4.9%, greater than about 4.8%, greater than about 4.7%, greater than about 4.6%, greater than about 4.5%, greater than about 4.4%, greater than about 4.3%, greater than about 4.2%, greater than about 4.1%, greater than about 4.0%, greater than about 3.9%, greater than about 3.8%, greater than about 3.7%, or greater than about 3.6%. 【0243】 In one embodiment, the compositions described herein comprise a total of risankizumab with M5, risankizumab with M6, and risankizumab with M7 in an amount that is about 5.3%, about 5.2%, about 5.1%, about 5.0%, about 4.9%, about 4.8%, about 4.7%, about 4.6%, about 4.5%, about 4.4%, about 4.3%, about 4.2%, about 4.1%, about 4.0%, about 3.9%, about 3.8%, about 3.7%, or about 3.6% of total risankizumab species with N-glycosylation. 【0244】 In one embodiment, the present disclosure relates to a composition comprising about 150 mg / ml of an antibody comprising a light chain having the amino acid sequence of SEQ ID NO: 9 and a heavy chain having the amino acid sequence of SEQ ID NO: 10, with or without N-glycosylation, wherein less than about 5.4% (e.g., about 3.6% to about 4.1%, about 4.3% to about 4.9%, or about 3.6% to about 4.9%) of all antibody species with N-glycosylation have high mannose N-glycans (e.g., M5, M6, and M7). 【0245】 In one embodiment, the high mannose N-glycan is M5. 【0246】 In one embodiment, the compositions described herein comprise multiple risankizumab species with or without N-glycosylation, and the level of risankizumab comprising M5 is less than 5.3% of the total risankizumab species with N-glycosylation. For example, and without limitation, the level of risankizumab comprising M5 in the compositions described herein may be less than about 5.2%, less than about 5.1%, less than about 5.0%, less than about 4.9%, less than about 4.8%, less than about 4.7%, less than about 4.6%, less than about 4.5%, less than about 4.4%, less than about 4.3%, less than about 4.2%, less than about 4.1%, less than about 4.0%, less than about 3.9%, less than about 3.8%, less than about 3.7%, less than about 3.6%, less than about 3.5%, less than about 3.4%, less than about 3.3%, less than about 3.2%, less than about 3.1%, or less than about 3. N-glycosylated risankizumab species are present in a detectable amount that is less than 0%, less than about 2.9%, or less than about 2.8%, or any range therebetween (inclusive), e.g., about 2.7% to 5.2%, about 3.1% to about 5.2%, about 3.5% to about 5.2%, about 4.0% to about 5.2%, about 4.5% to about 5.2%, about 5% to about 5.2%, about 2.7% to about 5.0%, about 2.7% to about 4.5%, about 2.7% to about 4.0%, about 2.7% to about 3.5%, about 2.7% to about 3.1%, about 3.2% to about 3.7%, or about 2.7% to about 3.7% of risankizumab species with N-glycosylation. 【0247】 In one embodiment, the compositions described herein comprise risankizumab containing a detectable amount of M5 that is less than about 5.3% of total risankizumab species with N-glycosylation but greater than about 5.2%, greater than about 5.1%, greater than about 5.0%, greater than about 4.9%, greater than about 4.8%, greater than about 4.7%, greater than about 4.6%, greater than about 4.5%, greater than about 4.4%, greater than about 4.3%, greater than about 4.2%, greater than about 4.1%, greater than about 4.0%, greater than about 3.9%, greater than about 3.8%, greater than about 3.7%, greater than about 3.6%, greater than about 3.5%, greater than about 3.4%, greater than about 3.3%, greater than about 3.2%, greater than about 3.1%, greater than about 3.0%, greater than about 2.9%, or greater than about 2.8%, or greater than about 2.7%. 【0248】 In one embodiment, the compositions described herein comprise risankizumab comprising M5 in an amount that is about 2.7%, about 2.8%, about 2.9%, about 3.0%, about 3.1%, about 3.2%, about 3.3%, about 3.4%, about 3.5%, about 3.6%, about 3.7%, about 3.8%, about 3.9%, about 4.0%, about 4.1%, about 4.2%, about 4.3%, about 4.4%, about 4.5%, about 4.6%, about 4.7%, about 4.8%, about 4.9%, about 5.0%, about 5.1%, or about 5.2% of total risankizumab species with N-glycosylation. 【0249】 In one embodiment, the disclosure relates to a composition comprising about 150 mg / ml of risankizumab, with or without N-glycosylation, wherein less than about 5.3% (e.g., about 2.7% to about 3.1%, about 3.2% to about 3.7%, or about 2.7% to about 3.7%) of all risankizumab species with N-glycosylation have M5. 【0250】 In one embodiment, the high mannose N-glycan is M6. 【0251】 In one embodiment, the compositions described herein comprise multiple risankizumab species with or without N-glycosylation, and the level of risankizumab comprising M6 is less than 2.6% of the total risankizumab species with N-glycosylation. For example, and without limitation, the level of risankizumab comprising M6 in the compositions described herein may be less than about 2.5%, less than about 2.4%, less than about 2.3%, less than about 2.2%, less than about 2.1%, less than about 2.0%, less than about 1.9%, less than about 1.8%, less than about 1.7%, less than about 1.6%, less than about 1.5%, less than about 1.4%, less than about 1.3%, less than about 1.2%, less than about 1.1%, less than about 1.0%, less than about 0.9%, less than about 0.8%, less than about 0.7%, less than about 0.6%, or less than about 0.5%, or any of its equivalents, of total risankizumab species with N-glycosylation. The N-glycosylated risankizumab species are present in a detectable amount in any range between (inclusive), for example, about 0.4% to about 2.5%, about 0.4% to about 2.4%, about 0.4% to about 2.2%, about 0.4% to about 2.0%, about 0.4% to about 1.8%, about 0.4% to about 1.6%, about 0.4% to about 1.4%, about 0.4% to about 1.2%, about 0.4% to about 1.0%, about 0.4% to about 0.9%, about 0.4% to about 0.8%, about 0.4% to about 0.7%, about 0.4% to about 0.6%, about 0.4% to about 0.5%, or about 0.6% to about 0.7% of total N-glycosylated risankizumab species. 【0252】 In one embodiment, the compositions described herein comprise risankizumab with a detectable amount of M6 that is less than about 2.6% of total risankizumab species with N-glycosylation, but is greater than about 2.5%, greater than about 2.4%, greater than about 2.3%, greater than about 2.2%, greater than about 2.1%, greater than about 2.0%, greater than about 1.9%, greater than about 1.8%, greater than about 1.7%, greater than about 1.6%, greater than about 1.5%, greater than about 1.4%, greater than about 1.3%, greater than about 1.2%, greater than about 1.1%, greater than about 1.0%, greater than about 0.9%, greater than about 0.8%, greater than about 0.7%, greater than about 0.6%, greater than about 0.5%, or greater than about 0.4%. 【0253】 In one embodiment, the compositions described herein comprise risankizumab containing M6 in an amount that is about 2.5%, about 2.4%, about 2.3%, about 2.2%, about 2.1%, about 2.0%, about 1.9%, about 1.8%, about 1.7%, about 1.6%, about 1.5%, about 1.4%, about 1.3%, about 1.2%, about 1.1%, about 1.0%, about 0.9%, about 0.8%, about 0.7%, about 0.6%, about 0.5%, or about 0.4% of total risankizumab species with N-glycosylation. 【0254】 In one embodiment, the disclosure relates to a composition comprising about 150 mg / ml of risankizumab, with or without N-glycosylation, wherein less than about 2.6% (e.g., about 0.4% to about 0.5%, about 0.6% to about 0.7%, or about 0.4% to about 0.7%) of all risankizumab species with N-glycosylation have M6. 【0255】 In one embodiment, the high mannose N-glycan is M7. 【0256】 In one embodiment, the compositions described herein include multiple risankizumab species with or without N-glycosylation, and the level of risankizumab comprising M7 is less than 2.0% of the total risankizumab species with N-glycosylation. For example, and without limitation, the level of risankizumab comprising M6 in the compositions described herein is less than about 1.9%, less than about 1.8%, less than about 1.7%, less than about 1.6%, less than about 1.5%, less than about 1.4%, less than about 1.3%, less than about 1.2%, less than about 1.1%, less than about 1.0%, less than about 0.9%, less than about 0.8%, less than about 0.7%, less than about 0.6%, less than about 0.5%, or more than about 0.4%, or any range therebetween. It is present in a detectable amount in any range (inclusive), e.g., about 0.4% to about 1.9%, about 0.4% to about 1.8%, about 0.4% to about 1.6%, about 0.4% to about 1.4%, about 0.4% to about 1.2%, about 0.4% to about 1.0%, about 0.4% to about 0.9%, about 0.4% to about 0.8%, about 0.4% to about 0.7%, about 0.4% to about 0.6%, about 0.4% to about 0.5%, or about 0.5% to about 0.6% of total N-glycosylated risankizumab species. 【0257】 In one embodiment, the compositions described herein comprise risankizumab containing a detectable amount of M7 that is less than about 2.0% of total risankizumab species with N-glycosylation, but greater than about 1.9%, greater than about 1.8%, greater than about 1.7%, greater than about 1.6%, greater than about 1.5%, greater than about 1.4%, greater than about 1.3%, greater than about 1.2%, greater than about 1.1%, greater than about 1.0%, greater than about 0.9%, greater than about 0.8%, greater than about 0.7%, greater than about 0.6%, greater than about 0.5%, or greater than about 0.4%. 【0258】 In one embodiment, the compositions described herein comprise risankizumab containing M7 in an amount that is about 1.9%, about 1.8%, about 1.7%, about 1.6%, about 1.5%, about 1.4%, about 1.3%, about 1.2%, about 1.1%, about 1.0%, about 0.9%, about 0.8%, about 0.7%, about 0.6%, about 0.5%, or about 0.4% of total risankizumab species with N-glycosylation. 【0259】 In one embodiment, the disclosure relates to a composition comprising about 150 mg / ml risankizumab, with or without N-glycosylation, wherein less than about 2.0% (e.g., about 0.4% to about 0.5%, about 0.5% to about 0.6%, or about 0.4% to about 0.6%) of all risankizumab species with N-glycosylation have M7. 【0260】 In one embodiment, the amount of risankizumab comprising high mannose N-glycans in the compositions described herein can be determined using methods known in the art, for example, by 2-AB and HILIC-FL chromatography (e.g., the 2-AB and HILIC-FL chromatography method described in Example 12), or RapiFluor HILIC-FL chromatography (using the RapiFluor HILIC-FL chromatography method described in Example 13). 【0261】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 185 mM trehalose, 10 mM acetate, and 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7, and wherein less than about 5.4% of all risankizumab species with N-glycosylation have high mannose N-glycans (e.g., M5, M6, and / or M7). 【0262】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7, and wherein less than about 5.4% of all risankizumab species with N-glycosylation have high mannose N-glycans (e.g., M5, M6, and / or M7). 【0263】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7, and wherein less than about 5.4% of total risankizumab species with N-glycosylation have high mannose N-glycans (e.g., M5, M6, and / or M7), and the amount of risankizumab that has high mannose N-glycans is determined by 2-AB and HILIC-FL chromatography (e.g., using the 2-AB and HILIC-FL chromatography method described in Example 12) or by RapiFluor HILIC-FL chromatography (e.g., using the RapiFluor HILIC-FL chromatography method described in Example 13). 【0264】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, and 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7, and the total amount of risankizumab containing M5, risankizumab containing M6, and risankizumab containing M7 is less than about 5.4% (e.g., about 3.6% to about 4.1%, about 4.3% to 4.9%, or about 3.6% to about 3.6%) of all risankizumab species with N-glycosylation. 【0265】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, and 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7; the sum of risankizumab with M5, risankizumab with M6, and risankizumab with M7 is less than about 5.4% (e.g., about 3.6% to about 4.1%, about 4.3% to 4.9%, or about 3.6% to about 4.9%) of total risankizumab species with N-glycosylation; and the amount of risankizumab containing high mannose N-glycans is determined by 2-AB and HILIC-FL chromatography (e.g., using the 2-AB and HILIC-FL chromatography methods described in Example 12) or RapiFluor 1000 (e.g., RapiFluor 1000) HPLC. As determined by HILIC-FL chromatography (eg, using the RapiFluor HILIC-FL chromatography method described in Example 13). 【0266】 Liquid formulations encompassed by the present disclosure may contain added water, such as USP grade water. 【0267】 In some embodiments, greater than about 84.4% of all N-glycosylated risankizumab species have complex fucosylated oligosaccharides. For example, but not limited to, the level of risankizumab containing complex fucosylated oligosaccharides in the compositions described herein is present in an amount that is greater than about 85%, greater than about 85.5%, greater than about 86%, greater than about 86.5%, greater than about 87%, greater than about 87.5%, greater than about 88%, greater than about 88.5%, greater than about 89%, greater than about 89.5%, greater than about 90%, or greater than about 90.5% of all N-glycosylated risankizumab species. 【0268】 In some embodiments, the level of risankizumab comprising complex fucosylated oligosaccharides in the compositions described herein is present in an amount that is about 85% to about 91%, about 88.0% to about 88.9%, about 89.8% to about 90.9%, or 88.0% to 90.9% of total risankizumab species with N-glycosylation. In some embodiments, the level of risankizumab comprising complex fucosylated oligosaccharides in the compositions described herein is present in an amount that is about 88.0%, about 88.3%, about 88.4%, about 88.9%, about 89.8%, about 90.2%, or about 90.9% of total risankizumab species with N-glycosylation. 【0269】 In some embodiments, the level of risankizumab containing complex fucosylated oligosaccharides is determined by 2-AB and HILIC-FL chromatography or by RapiFluor HILIC-FL chromatography. 【0270】 In some embodiments, the composition comprises about 0.8% to about 1.4% (e.g., about 0.8%, about 0.9%, about 1.0%, about 1.1%, about 1.2%, about 1.3%, or about 1.4%) aglycosylated risankizumab. 【0271】 In some embodiments, the aglycosylated risankizumab is determined by tryptic peptide mapping (e.g., using the tryptic peptide mapping analysis described in Example 14). 【0272】 b. Risankizumab compositions with improved purity In one embodiment, the compositions provided herein have at least characteristic (b) that at least about 99.1% of the risankizumab is present as a monomer and / or about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by ultra-performance size exclusion chromatography (UP-SEC). For example, the compositions described herein comprise risankizumab, and at least about 99.1% of the risankizumab is present as a monomer and about 0.4% or less of the risankizumab is present as a high molecular weight (HMW) species, as measured by UP-SEC. 【0273】 In one embodiment, at least about 99.1% of the risankizumab is present as a monomer in the compositions provided herein as measured by UP-SEC. For example, but not limited to, at least about 99.2%, at least about 99.3%, at least about 99.4%, at least about 99.5%, at least about 99.6%, or at least about 99.7% of the risankizumab is present as a monomer as measured by UP-SEC, or any range therebetween (inclusive), such as about 99.1% to about 99.7%, 99.1% to about 99.6%, about 99.2% to about 99.7%, or about 99.2% to about 99.6% of the risankizumab is present as a monomer as measured by UP-SEC. 【0274】 In one embodiment, when measured by UP-SEC, about 0.4% or less of risankizumab is present as a high molecular weight (HMW) species in the compositions provided herein. For example, but not limited to, when measured by UP-SEC, about 0.35% or less, about 0.3% or less, about 0.25% or less, about 0.2% or less, about 0.15% or less, or about 0.1% or less of risankizumab is present as a high molecular weight (HMW) species, or any range therebetween (inclusive), when measured by UP-SEC, such as about 0.1% to about 0.4%, about 0.1 to about 0.3%, about 0.1 to about 0.2%, about 0.2% to about 0.4%, or about 0.2% to about 0.3% of risankizumab is present as a high molecular weight (HMW) species. 【0275】 In one embodiment, the disclosure relates to a composition comprising about 150 mg / ml risankizumab, wherein at least about 99.1% (about 99.1% to about 99.7%, 99.1% to about 99.6%, about 99.2% to about 99.7%, or about 99.2% to about 99.6%) of the risankizumab is present as monomers and / or about 0.4% or less (about 0.1% to about 0.4%, about 0.1 to about 0.3%, about 0.1 to about 0.2%, about 0.2% to about 0.4%, or about 0.2% to about 0.3%) of the risankizumab is present as high molecular weight (HMW) species, as measured by ultra-performance size exclusion chromatography (UP-SEC). 【0276】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 185 mM trehalose, 10 mM acetate, and 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7 and a CHO of at least about 99.1% (about 99.1% to about 99.7%, 99.1% to about 99.6%, about 99.2% to about 99.7%, or about 99.2% to about 99.6%). Risankizumab is present as a monomer and / or about 0.4% or less (about 0.1% to about 0.4%, about 0.1 to about 0.3%, about 0.1 to about 0.2%, about 0.2% to about 0.4%, or about 0.2% to about 0.3%) of risankizumab is present as a high molecular weight (HMW) species when measured by ultra-performance size exclusion chromatography (UP-SEC) (e.g., using the UP-SEC method described in Example 15). 【0277】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7 and a solubility of at least about 99.1% (about 99.1% to about 99.7%, 99.1% to about 99.6%, about 99.2% to about 99.7%, or About 99.2% to about 99.6% of the risankizumab is present as monomers, and / or about 0.4% or less (about 0.1% to about 0.4%, about 0.1 to about 0.3%, about 0.1 to about 0.2%, about 0.2% to about 0.4%, or about 0.2% to about 0.3%) of the risankizumab is present as high molecular weight (HMW) species when measured by ultra-performance size-exclusion chromatography (UP-SEC) (e.g., using the UP-SEC method described in Example 15). 【0278】 Liquid formulations encompassed by the present disclosure may contain added water, such as USP grade water. 【0279】 In one embodiment, the compositions described herein have at least characteristic (c) that, when measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR), greater than about 97.5% of the risankizumab is present as a main peak and / or less than about 2.2% of the risankizumab is present as low molecular weight (LMW) species. For example, the compositions described herein comprise risankizumab, and, when measured by CGE-NR, greater than about 97.5% of the risankizumab is present as a main peak and less than about 2.2% of the risankizumab is present as low molecular weight (LMW) species. 【0280】 In one embodiment, greater than about 97.5% of the risankizumab is present as the main peak when measured by CGE-NR. For example, but not limited to, greater than about 97.6%, greater than about 97.7%, greater than about 97.8%, greater than about 97.9%, greater than about 98.0%, greater than about 98.1%, greater than about 98.2%, greater than about 98.3%, or greater than about 98.4% of the risankizumab is present as the main peak when measured by CGE-NR, or any range therebetween (both endpoints) when measured by CGE-NR. The main peak is present in an amount of, for example, about 97.6% to about 98.4%, about 97.6% to about 98.3%, about 97.6% to about 98.2%, about 97.7% to about 98.4%, about 97.7% to about 98.3%, about 97.7% to about 98.2%, about 97.8% to about 98.4%, about 97.8% to about 98.3%, or about 97.8% to about 98.2% risankizumab. 【0281】 In one embodiment, when measured by CGE-NR, less than about 2.2% of risankizumab is present as a low molecular weight (LMW) species. For example, but not limited to, less than about 2.1%, less than 2.0%, less than 1.9%, less than 1.8%, less than 1.7%, less than 1.6%, or less than 1.5% of risankizumab is present as a low molecular weight (LMW) species, or any range therebetween (inclusive), when measured by CGE-NR, such as about 1.5% to about 2.1%, about 1.6% to about 2.1%, about 1.7% to about 2.1%, about 1.5% to about 2.0%, about 1.6% to about 2.0%, or about 1.7% to about 2.0% of risankizumab is present as a low molecular weight (LMW) species. 【0282】 In one embodiment, the present disclosure relates to a composition comprising about 150 mg / ml risankizumab, having a saturation of greater than about 97.5% (e.g., about 97.6% to about 98.4%, about 97.6% to about 98.3%, about 97.6% to about 98.2%, about 97.7% to about 98.4%, about 97.7% to about 98.3%, about 97.7% to about 98.2%, about 97.8% to about 98.4%, about 97.8% to about 98.3%, or about 97.8% to about 98.2%). ) risankizumab is present as the main peak, and / or less than about 2.2% (e.g., about 1.5% to about 2.1%, about 1.6% to about 2.1%, about 1.7% to about 2.1%, about 1.5% to about 2.0%, about 1.6% to about 2.0%, or about 1.7% to about 2.0%) of risankizumab is present as a low molecular weight (LMW) species as measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR). 【0283】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 185 mM trehalose, 10 mM acetic acid, and 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7 and a CHCl3 content of greater than about 97.5% (e.g., about 97.6% to about 98.4%, about 97.6% to about 98.3%, about 97.6% to about 98.2%, about 97.7% to about 98.4%, about 97.7% to about 98.3%, about 97.7% to about 98.2%, about 97.8% to about 98.4%, about 97.8% to about 98.5%). 0.3%, or about 97.8% to about 98.2%, of the risankizumab is present as a main peak, and / or less than about 2.2% (e.g., about 1.5% to about 2.1%, about 1.6% to about 2.1%, about 1.7% to about 2.1%, about 1.5% to about 2.0%, about 1.6% to about 2.0%, or about 1.7% to about 2.0%) of the risankizumab is present as a low molecular weight (LMW) species when measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR) (e.g., using the CGE-NR method described in Example 15). 【0284】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, and the pharmaceutical composition has a pH of about 5.7 and a CHCl3 content of greater than about 97.5% (e.g., about 97.6% to about 98.4%, about 97.6% to about 98.3%, about 97.6% to about 98.2%, about 97.7% to about 98.4%, about 97.7% to about 98.3%, about 97.7% to about 98.2%, about 97.8% to about 98.9%). About 98.4%, about 97.8% to about 98.3%, or about 97.8% to about 98.2% of the risankizumab is present as a main peak, and / or less than about 2.2% (e.g., about 1.5% to about 2.1%, about 1.6% to about 2.1%, about 1.7% to about 2.1%, about 1.5% to about 2.0%, about 1.6% to about 2.0%, or about 1.7% to about 2.0%) of the risankizumab is present as a low molecular weight (LMW) species when measured by capillary gel electrophoresis under non-reducing conditions (CGE-NR) (e.g., using the CGE-NR method described in Example 15). 【0285】 Liquid formulations encompassed by the present disclosure may contain added water, such as USP grade water. 【0286】 c. Risankizumab Compositions with Reduced Immunogenicity In one embodiment, the compositions described herein have at least the characteristic (d) that the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% following administration of a single subcutaneous dose of the pharmaceutical composition to a human. 【0287】 For example, and without limitation, the incidence of treatment-emergent ADAs is less than about 4.5%, less than about 4.0%, less than about 3.5%, less than about 3.0%, less than about 2.5%, less than about 2.0%, less than about 1.5%, less than about 1.0%, less than about 0.5%, less than about 0.4%, less than about 0.3%, less than about 0.2%, less than about 0.1%, less than about 0.01%, less than about 0.001%, or less than about 0.0001%. 【0288】 In one embodiment, the incidence of treatment-emergent ADA is about 0.0%. 【0289】 In one embodiment, the incidence of treatment-emergent ADA is measured after humans receive a single subcutaneous injection of a 150 mg dose of the pharmaceutical composition. 【0290】 In one embodiment, the presence of ADA is determined by using a validated bridging electrochemiluminescence immunoassay. 【0291】 In one embodiment, the incidence of treatment-emergent ADA is less than about 4.7%, e.g., about 0.0%, after administration of a single subcutaneous injection of a 150 mg dose of the pharmaceutical composition, as measured using a bridging electrochemiluminescence immunoassay (e.g., using the bridging electrochemiluminescence immunoassay described in Example 16). 【0292】 In one embodiment, the disclosure relates to a composition comprising about 150 mg / ml risankizumab, wherein the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% (e.g., about 0.0%) after administration of a single subcutaneous injection of a 150 mg dose of the pharmaceutical composition to a human. 【0293】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 185 mM trehalose, 10 mM acetate, and 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7, and the incidence of treatment-emergent anti-drug antibodies (ADA) is less than about 4.7% (e.g., about 0.0%) after administration of a single subcutaneous injection of a 150 mg dose of the pharmaceutical composition to humans. 【0294】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 185 mM trehalose, 10 mM acetate, and 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7 and has an incidence of treatment-emergent anti-drug antibodies (ADA) of less than about 4.7% (e.g., about 0.0%) after administration of a single subcutaneous injection of a 150 mg dose of the pharmaceutical composition, as determined by using a bridging electrochemiluminescence immunoassay (e.g., by using the bridging electrochemiluminescence immunoassay described in Example 16). 【0295】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7 and has an incidence of treatment-emergent anti-drug antibodies (ADA) of less than about 4.7% (e.g., about 0.0%) following administration of a single subcutaneous injection of a 150 mg dose of the pharmaceutical composition to humans. 【0296】 In one embodiment, a pharmaceutical composition described herein comprises about 150 mg / mL risankizumab, 0.054 mg / mL acetic acid, 1.24 mg / mL sodium acetate trihydrate, 70 mg / mL trehalose dihydrate, 0.20 mg / mL polysorbate 20, wherein the pharmaceutical composition has a pH of about 5.7 and has an incidence of treatment-emergent anti-drug antibodies (ADA) of less than about 4.7% (e.g., about 0.0%) after administration of a single subcutaneous injection of a 150 mg dose of the pharmaceutical composition to humans, as determined by using a bridging electrochemiluminescence immunoassay (e.g., by using the bridging electrochemiluminescence immunoassay described in Example 16). 【0297】 Liquid formulations encompassed by the present disclosure may contain added water, such as USP grade water. 【0298】 Risankizumab Compositions Containing Poloxamer 188 In one aspect, the disclosure relates to a composition comprising (1) risankizumab and (2) poloxamer 188 (P188), wherein the composition does not include polysorbate 20 (PS20) and / or polysorbate 80 (PS80). 【0299】 In one embodiment, the compositions described herein contain about 60 mg / ml to about 150 mg / ml of risankizumab. For example, but not limited to, the compositions described herein may contain about 70 mg / ml to about 150 mg / ml, about 80 mg / ml to about 150 mg / ml, about 90 mg / ml to about 150 mg / ml, about 100 mg / ml to about 150 mg / ml, about 110 mg / ml to about 150 mg / ml, about 120 mg / ml to about 150 mg / ml, about 130 mg / ml to about 150 mg / ml, or about 140 mg / ml to about 150 mg / ml. , 60 mg / ml to about 70 mg / ml, 60 mg / ml to about 80 mg / ml, 60 mg / ml to about 90 mg / ml, 60 mg / ml to about 100 mg / ml, 60 mg / ml to about 110 mg / ml, 60 mg / ml to about 120 mg / ml, 60 mg / ml to about 130 mg / ml, or 60 mg / ml to about 140 mg / ml of risankizumab, and ranges and amounts between any of these aforementioned concentrations. 【0300】 In one embodiment, the compositions described herein contain about 60 mg / ml, about 70 mg / ml, about 80 mg / ml, about 90 mg / ml, about 100 mg / ml, about 110 mg / ml, about 120 mg / ml, about 130 mg / ml, about 140 mg / ml, or about 150 mg / ml of risankizumab. 【0301】 In one embodiment, the compositions described herein further comprise phospholipase A2 (PLA2) in an amount that is greater than about 250 pg per mg of risankizumab. 【0302】 In one embodiment, the compositions described herein contain more than about 260 pg, more than about 270 pg, more than about 280 pg, more than about 290 pg, more than about 300 pg, more than about 310 pg, more than about 320 pg, more than about 330 pg, more than about 340 pg, more than about 350 pg, more than about 360 pg, more than about 380 pg, more than about 400 pg, more than about 450 pg, more than about 500 pg, more than about 550 pg, more than about 560 pg, more than about 570 pg, more than about 580 pg, more than about 590 pg, more than about 600 pg, more than about 610 pg, more than about 620 pg, more than about 630 pg, more than about 640 pg, more than about 650 pg, more than about 660 pg, more than about 670 pg, more than about 680 pg, more than about 690 pg, more than about 700 pg, more than about 710 pg, more than about 720 pg, more than about 750 pg, more than about 760 pg, more than about 770 pg, more than about 780 pg, more than about 790 pg, more than about 800 pg, more than about 810 pg, more than about 820 pg, more than about 830 pg, more than about 840 pg, more than about 850 pg, more than about 860 pg, more than about 870 pg, more than about 880 pg, more than about 890 pg, more than about 900 pg, more than about 910 pg, more than g, greater than about 500 pg, greater than about 550 pg, greater than about 600 pg, greater than about 650 pg, greater than about 700 pg, greater than about 750 pg, greater than about 800 pg, greater than about 900 pg, or about 1000 pg, or any range therebetween (inclusive), e.g., about 250 pg to about 1100 pg, about 260 pg to about 1100 pg per mg of risankizumab , about 270pg to about 1100pg, about 280pg to about 1100pg, about 290pg to about 1100pg, about 300pg to about 1100pg, about 310pg to about 1100 pg, about 320pg to about 1100pg, about 340pg to about 1100pg, about 360pg to about 1100pg, about 250pg to about 1000pg, about 250pg to about 90 The amount of PLA2 is 0 pg, about 250 pg to about 800 pg, about 250 pg to about 700 pg, about 250 pg to about 600 pg, about 250 pg to about 500 pg, about 250 pg to about 400 pg, about 250 pg to about 1030 pg, about 290 pg to about 1090 pg, about 360 pg to about 450 pg, or about 310 pg to about 920 pg. 【0303】 In one embodiment, the compositions described herein comprise PLA2 in an amount that is about 260 pg, about 270 pg, about 280 pg, about 290 pg, about 300 pg, about 310 pg, about 320 pg, about 330 pg, about 340 pg, about 350 pg, about 360 pg, about 380 pg, about 400 pg, about 450 pg, about 500 pg, about 550 pg, about 600 pg, about 650 pg, about 700 pg, about 750 pg, about 800 pg, about 900 pg, about 1000 pg, or about 1100 pg per mg of risankizumab. 【0304】 The PLA2 in the compositions described herein can be PLA2G2, PLA2G15, or a combination thereof. In one embodiment, the PLA2 is PLA2G15. 【0305】 The amount of PLA2 in the compositions described herein can be determined by methods known in the art, for example, by mass spectrometry or by ELISA.In one embodiment, the amount of PLA2 in the compositions described herein is determined by ELISA, for example, the ELISA method described in Example 9. 【0306】 In some embodiments, the composition does not include PS80. 【0307】 Risankizumab purification In one embodiment, risankizumab can be recombinantly produced in a variety of host cells using methods described in the Examples (e.g., Example 3) or using methods known in the art, such as using cell culture media using hydrolysate-based media or chemically defined media containing specific ranges of manganese and / or galactose (see, e.g., U.S. Pat. No. 9,062,106), or by using recombinant host cells overexpressing β1,4 galactosyltransferase, or using host cells with beta-galactosidase knockdown (U.S. Pat. No. 9,550,826). U.S. Pat. Nos. 9,062,106 and 9,550,826 are incorporated herein by reference in their entireties. 【0308】 The risankizumab compositions described herein can be produced using the exemplary optimized purification process described in Example 3 and FIG. 14 herein, and are illustrated below. 【0309】 Once a clarified solution or mixture containing antibodies is obtained, separation of the antibodies from other proteins produced by cells such as HPs may be performed using a combination of different purification techniques, including, but not limited to, affinity separation, ion exchange separation, mixed-mode separation, and hydrophobic interaction separation (alone or in combination). The separation steps separate the protein mixture based on biophysical properties such as charge, degree of hydrophobicity, and / or size, depending on the particular form of separation, including, but not limited to, chromatographic separation. In one embodiment of the present disclosure, separation may be performed using chromatography, including, but not limited to, cationic, anionic, hydrophobic interaction, and / or mixed-mode chromatography. For each of these techniques, several different chromatographic resins are commercially available, allowing the purification scheme to be precisely tailored to the specific proteins involved. The essence of each of the separation methods described is to move proteins at different speeds through a chromatographic medium, such as a resin in a column, to achieve physical separation that increases with further passage through the chromatographic medium, or to selectively attach proteins to a chromatographic medium, such as a separation resin in a column, followed by differential elution using different eluents. In some cases, the antibody is separated from the HP when the HP specifically adheres to the chromatographic medium, such as the resin of a column, but the antibody does not, i.e., the antibody is contained in the eluent, while in other cases the antibody of interest adheres to the chromatographic medium, such as the resin of a column, while the HP is pushed off the column during a wash cycle. 【0310】 a. Primary collection 【0311】 In certain embodiments, it may be advantageous to subject samples produced in accordance with the present disclosure to at least a first stage of clarification and primary recovery. 【0312】 Primary recovery can include one or more centrifugation steps to further clarify the sample mixture and thereby aid in the purification of the protein of interest. Centrifugation of the sample can be performed, for example, but not limited to, at 7,000 x g to about 12,750 x g. In the context of large-scale purification, such centrifugation can be performed online, for example, but not limited to, at a flow rate set to achieve a turbidity level of 150 NTU in the resulting supernatant. Such supernatant can then be collected for further purification. 【0313】 In certain embodiments, primary recovery may also include the use of one or more depth filtration steps to further clarify the sample matrix and thereby aid in the purification of antibodies produced using the cell culture techniques of the present disclosure. Depth filters include filter media with graded densities. Such graded densities allow larger particles to be trapped near the surface of the filter, while smaller particles penetrate the larger open areas of the filter surface and are trapped only in the smaller openings closer to the center of the filter. In certain embodiments, the depth filtration step may be a delipidation depth filtration step. While certain embodiments use a depth filtration step only during the primary recovery stage, other embodiments may use depth filters, including delipidation depth filters, during one or more additional purification stages. Non-limiting examples of depth filters that may be used in the context of the present disclosure include X0HC depth filters, DOHC depth filters, Cuno™ Model 30 / 60ZA depth filters (3M Corp.), and 0.45 / 0.2 μm Sartopore™ dual-layer filter cartridges. 【0314】 B. Affinity chromatography 【0315】 In certain embodiments, it may be advantageous to subject risankizumab produced according to the present disclosure to affinity chromatography to further purify the antibody from HP (e.g., lipase). In certain embodiments, the chromatography material may be capable of selectively or specifically binding to risankizumab. Non-limiting examples of such chromatography materials include Protein A, Protein G, chromatography materials containing, for example, an antigen bound by the antibody of interest, and chromatography materials containing an Fc-binding protein. In some embodiments, the affinity chromatography step may involve subjecting the primary recovery sample to a column containing a suitable Protein A resin. In certain embodiments, Protein A resin may be useful for affinity purification and isolation of various antibody isotypes, particularly IgG1, IgG2, and IgG4. Protein A is a bacterial cell wall protein that binds mammalian IgG primarily via its Fc region. In its native state, Protein A has five IgG-binding domains and other domains of unknown function. 【0316】 There are several commercial sources of Protein A resin. One suitable resin can be MabSelect™ from GE Healthcare. Another suitable resin can be MabSelect SuRe™. A non-limiting example of a suitable column packed with MabSelect™ is a column approximately 1.0 cm in diameter by approximately 21.6 cm in length (approximately 17 ml bed volume). A column of this size can be used for small-scale purification and can be compared to other columns used for scale-up. For example, a 20 cm x 21 cm column with a bed volume of approximately 6.6 L can be used for larger-scale purification. Regardless of the column, the column can be packed using a suitable resin, such as MabSelect™ or MabSelect SuRe™. 【0317】 C. Ion-exchange chromatography 【0318】 In certain embodiments, it may be advantageous to subject risankizumab produced according to the present disclosure to ion exchange chromatography to purify risankizumab from HP (e.g., lipase). Ion exchange separation includes any method that separates two substances based on the difference in their ionic charge, and may use either a cation exchange material or an anion exchange material. For example, the use of a cation exchange material over an anion exchange material is based on the localized charge of the protein. Thus, the use of an anion exchange step before a cation exchange step, or the use of a cation exchange step before an anion exchange step, is encompassed by the present disclosure. Furthermore, the use of only a cation exchange step, only an anion exchange step, or any sequential combination of the two is encompassed by the present disclosure. 【0319】 In carrying out the separation, the initial protein mixture can be contacted with an ion exchange material using any of a variety of techniques, for example, using batch purification techniques or chromatographic techniques. 【0320】 Anionic or cationic substituents may be attached to the matrix to form anionic or cationic supports for chromatography. Non-limiting examples of anionic exchange substituents include diethylaminoethyl (DEAE), quaternary aminoethyl (QAE), and quaternary amine (Q) groups. Cationic substituents include carboxymethyl (CM), sulfoethyl (SE), sulfopropyl (SP), phosphate (P), and sulfonate (S). Cellulose ion exchange resins such as DE23™, DE32™, DE52™, CM-23™, CM-32™, and CM-52™ are available from Whatman Ltd., Maidstone, Kent, UK. SEPHADEX®-based and -crosslinked ion exchangers are also known. For example, DEAE-, QAE-, CM-, and SP-SEPHADEX®, as well as DEAE-, Q-, CM-, and S-SEPHAROSE® and SEPHAROSE® Fast Fe, are all available from Pharmacia AB. Additionally, both DEAE- and CM-derivatized ethylene glycol-methacrylate copolymers, such as TOYOPEARL™ DEAE-6505 or M and TOYOPEARL™ CM-650S or M, are available from Toso Haas Co., Philadelphia, Pa. In some embodiments, cation exchange chromatography using Poros™ XS resin is used. 【0321】 d. Ultrafiltration / dialysis 【0322】 In certain embodiments, it may be advantageous to subject risankizumab produced according to the present disclosure to ultrafiltration and / or dialysis to purify the risankizumab from HPs (e.g., lipases). Ultrafiltration is described in detail in "Microfiltration and Ultrafiltration: Principles and Applications," L. Zeman and A. Zydney (Marcel Dekker, Inc., New York, NY, 1996), and "Ultrafiltration Handbook," Munir Cheryan (Technomic Publishing, 1986; ISBN No. 87762-456-9). One filtration process is tangential flow filtration, described in the Millipore catalog entitled "Pharmaceutical Process Filtration Catalogue," pp. 177-202 (Bedford, Mass., 1995 / 96). Ultrafiltration is generally considered to refer to filtration using filters with a pore size of less than 0.1 μm. By using a filter with such small pore size, the sample volume can be reduced by allowing the sample buffer to pass through the filter while the antibody is retained behind the filter. 【0323】 Dialysis is a method that uses an ultrafilter to remove and exchange salts, sugars, and non-aqueous solvents, separate them from bound species, remove low molecular weight materials, and / or cause a rapid change in the ionic and / or pH environment. Small solutes are most efficiently removed by adding solvent to the solution being ultrafiltered at a rate approximately equal to the ultrafiltration rate. This consistently washes minor species from the solution and effectively purifies retained proteins. In certain embodiments of the present disclosure, a dialysis step can be used to exchange various buffers used in connection with the present disclosure as well as to remove impurities from protein preparations, optionally prior to further chromatography or other purification steps. 【0324】 e. Hydrophobic interaction chromatography 【0325】 In certain embodiments, it may be advantageous to subject risankizumab produced according to the present disclosure to hydrophobic interaction chromatography to purify risankizumab from HP (lipase). For example, a first eluate obtained from an ion exchange column can be subjected to a hydrophobic interaction material to obtain a second eluate having a reduced HP level. A hydrophobic interaction chromatography (HIC) step as disclosed herein is generally performed to purify proteins, including removing HP. 【0326】 In performing HIC-based separations, the sample mixture is contacted with the HIC material, for example, using batch purification techniques or using a column. Prior to HIC purification, it may be desirable to remove any chaotropic agents or very hydrophobic substances, for example, by passing the mixture through a precolumn. 【0327】 While ion exchange chromatography relies on the charge of proteins to isolate them, hydrophobic interaction chromatography exploits the hydrophobicity of proteins. Hydrophobic groups on proteins interact with hydrophobic groups on the column. The more hydrophobic a protein is, the stronger its interaction with the column. Thus, the HIC step removes host cell-derived impurities (e.g., DNA and other high- and low-molecular-weight product-related species). 【0328】 HIC columns typically contain a base matrix (e.g., cross-linked agarose or synthetic copolymer materials) to which hydrophobic ligands (e.g., alkyl or aryl groups) are attached. Suitable HIC columns contain phenyl-substituted agarose resins (e.g., Phenyl Sepharose™ columns). Many HIC columns are commercially available. Examples include, but are not limited to, Phenyl Sepharose™ 6 Fast Flow columns with low or high substitution (Pharmacia LKB Biotechnology, AB, Sweden), phenyl Sepharose™ High Performance columns (Pharmacia LKB Biotechnology, AB, Sweden), Octyl Sepharose™ High Performance columns (Pharmacia LKB Biotechnology, AB, Sweden), Fractogel™ EMD Propyl or Fractogel™ EMD Phenyl columns (E. Merck, Germany), Macro-Prep™ Mehyl or Macro-Prep™ t-Butyl supports (Bio-Rad, California), WP HI-Propyl (C3)™ columns (JT Baker, New Jersey), and Toyopearl™ ether, phenyl, or butyl columns (TosoHaas, Pa.). 【0329】 f. Multimodal (mixed-mode) chromatography 【0330】 In certain embodiments, it may be advantageous to subject risankizumab produced according to the present disclosure to multimodal chromatography to purify risankizumab from HP (e.g., lipase). Multimodal chromatography is chromatography that utilizes a multimodal media resin. Such resins contain multimodal chromatography ligands. In certain embodiments, such ligands refer to ligands that can provide at least two distinct but cooperative sites for interacting with a substance to be bound. One of these sites provides an attractive type of charge-charge interaction between the ligand and the substance of interest. The other sites typically provide electron acceptor-donor interactions and / or hydrophobic and / or hydrophilic interactions. Electron donor-acceptor interactions include hydrogen bonding, π-π, cation-π, charge transfer, dipole-dipole, induced dipole, and other interactions. Multimodal chromatography ligands are also known as "mixed-mode" chromatography ligands. 【0331】 In certain embodiments, multimodal chromatography resins may be composed of multimodal ligands bound directly or via spacers to an organic or inorganic support, sometimes referred to as the base matrix. The support may be in the form of particles, such as essentially spherical particles, monoliths, filters, membranes, surfaces, capillaries, etc. In certain embodiments, the support may be prepared from natural polymers such as cross-linked carbohydrate materials, e.g., agarose, agar, cellulose, dextran, chitosan, konjac, carrageenan, gellan, alginic acid, etc. To achieve high adsorption capacity, the support may be porous, and the ligands are then bound to the exterior and pore surfaces. Such natural polymer supports can be prepared according to standard methods, such as inverse suspension gelation (S Hjerten: Biochim Biophys Acta 79(2), 393-398 (1964). Alternatively, the supports can be prepared from synthetic polymers, for example cross-linked synthetic polymers, such as styrene or styrene derivatives, divinylbenzene, acrylamide, acrylic acid esters, methacrylic acid esters, vinyl esters, vinylamides, etc. Such synthetic polymers can be produced according to standard methods, see for example "Styrene based polymer supports developed by suspension polymerization" (R ArShady: Chimica e L'Industria 70(9), 70-75 (1988)). Porous natural or synthetic polymer supports can be prepared according to standard methods, see for example Amersham Mixed-mode chromatography is also available from commercial sources such as Capto™ Adhere mixed-mode chromatography, which is available from Capto Biosciences, Uppsala, Sweden. In some embodiments, mixed-mode chromatography combines anion exchange (AEX) and hydrophobic interaction (HIC) functions. One non-limiting example of such mixed-mode chromatography suitable for the present disclosure is Capto™ Adhere mixed-mode chromatography. 【0332】 Characterization of risankizumab compositions Reducing hitchhiker proteins from risankizumab formulations beneficially increases the stability of the formulation (e.g., reduces particle formation, increases the shelf life of risankizumab pharmaceuticals, etc.). In one embodiment, no visible or shiny particles are observed in the liquid risankizumab compositions described herein for at least 3 months (e.g., at least 6 months, at least 9 months, at least 12 months, at least 15 months, at least 18 months, at least 21 months, at least 24 months, at least 27 months, at least 30 months, at least 33 months, or at least 36 months) at 2°C to 40°C (e.g., 4°C to 35°C, 4°C to 25°C, 4°C to 15°C, 4°C to 10°C, 2°C to 8°C, or any temperature within the above ranges, e.g., about 2°C, about 4°C, about 5°C, about 8°C, about 25°C, about 40°C, etc.). In one embodiment, no visible or shiny particles are observed in the liquid risankizumab compositions described herein for 24 months at about 4°C. 【0333】 In some embodiments, the liquid risankizumab compositions described herein comprise a surfactant with increased stability, which may be selected from the group consisting of polysorbate 20 (PS20), polysorbate 80 (PS80), polysorbate 40 (PS40), polysorbate 60 (PS60), polysorbate 65 (PS65), and poloxamer 188. 【0334】 The stability of a surfactant in a liquid risankizumab composition described herein can be assessed by directly measuring the amount of surfactant in the liquid risankizumab composition after storage at a particular temperature (e.g., 2°C, 4°C, 5°C, 8°C, 10°C, 12°C, 25°C, 30°C, 35°C, or 40°C) for a period of time (e.g., 3 months, 6 months, 9 months, 12 months, 18 months, 24 months, 30 months, 36 months, etc.). Alternatively, the stability of a surfactant in a liquid risankizumab composition described herein can be assessed by measuring the amount of degradation products of the surfactant (e.g., the amount of free fatty acids) in the risankizumab composition after storage at a particular temperature (e.g., 2°C, 4°C, 5°C, 8°C, 10°C, 12°C, 25°C, 30°C, 35°C, or 40°C) for a period of time (e.g., 3 months, 6 months, 9 months, 12 months, 18 months, 24 months, 30 months, 36 months, etc.). In some embodiments, the period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, or more months, or any range between 3 and 36 months, 12 and 24 months inclusive, etc. 【0335】 In some embodiments, the liquid risankizumab compositions described herein include PS20 at a concentration of, for example, 0.20 mg / mL, increasing the stability of PS20 in such liquid risankizumab compositions. 【0336】 In one embodiment, the stability of PS20 can be assessed by directly measuring the amount of PS20 in a risankizumab composition after storage for a period of time (e.g., 3 months, 6 months, 9 months, 12 months, 18 months, 24 months, 30 months, 36 months, etc.) at a particular temperature (e.g., 2°C, 4°C, 5°C, 8°C, 10°C, 12°C, 25°C, 30°C, 35°C, or 40°C). The amount of PS20 can be measured using any method known in the art, for example, using a high performance liquid chromatography-charged aerosol detector (HPLC-CAD), for example, the HPLC-CAD described in Example 10. In some embodiments, the period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, or more months, or any range between 3 and 36 months, 12 and 24 months inclusive, etc. 【0337】 For example, in some embodiments, at least 80% (e.g., at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of PS20 is retained after 6 months of storage at 5°C. In some embodiments, at least 70% (e.g., at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of PS20 is retained after 24 months of storage at 5°C. In some embodiments, at least 60% (e.g., at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of PS20 is retained after 6 months of storage at 25°C. In some embodiments, at least 40% (e.g., at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of PS20 is retained after 6 months of storage at 40°C. 【0338】 In one embodiment, the stability of PS20 can be assessed by measuring the amount of free fatty acids (FFAs), which are degradation products of PS20, in a risankizumab composition formulated with PS20 after storage at a particular temperature (e.g., 2°C, 4°C, 5°C, 8°C, 10°C, 12°C, 25°C, 30°C, 35°C, or 40°C) for a period of time (e.g., 3 months, 6 months, 9 months, 12 months, 18 months, 24 months, 30 months, 36 months, etc.). The amount of FFAs can be measured using any method known in the art, for example, using an enzymatic FFA or LC-FFA assay, such as the reverse-phase high-performance liquid chromatography UV (RP-HPLC-UV) method described in Example 10. In some embodiments, the period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, or more months, or any range between 3 and 36 months, 12 and 24 months inclusive, etc. 【0339】 For example, in some embodiments, the total amount of FFAs in a liquid composition formulated with PS20 described herein increases by 1.75-fold or less (e.g., 1.5-fold or less, 1.25-fold or less, or 1.1-fold or less) or does not increase after 6 months of storage at 5° C. In some embodiments, the total amount of FFAs in a liquid composition formulated with PS20 described herein is 20 nmol / ml or less (e.g., 18 nmol / ml or less, 15 nmol / ml or less, 12 nmol / ml or less, 10 nmol / ml or less, 8 nmol / ml or less, or 5 nmol / ml or less, or any range between 5 nmol / ml and 10 nmol / ml, etc., inclusive) after 6 months of storage at 5° C. In some embodiments, the FFAs are present in an amount below or at the detection limit of an FFA detection assay. 【0340】 In some embodiments, the total amount of FFAs in a liquid composition formulated with PS20 described herein increases by 3.5-fold or less (e.g., 3.2-fold or less, 3.0-fold or less, 2.5-fold or less, 2.0-fold or less, 1.8-fold or less, 1.6-fold or less, 1.4-fold or less, 1.2-fold or less, or 1.1-fold or less) or does not increase after 6 months of storage at 25° C. In some embodiments, the total amount of FFAs in a liquid composition formulated with PS20 described herein is 25 nmol / ml or less (e.g., 20 nmol / ml or less, 18 nmol / ml or less, 15 nmol / ml or less, 12 nmol / ml or less, 10 nmol / ml or less, 8 nmol / ml or less, or 5 nmol / ml or less, or any range between 5 nmol / ml and 10 nmol / ml, etc., inclusive) after 6 months of storage at 25° C. In some embodiments, the FFAs are present in an amount below or at the detection limit of an FFA detection assay. 【0341】 In some embodiments, the total amount of FFAs in a liquid composition formulated with PS20 described herein increases by 3-fold or less (e.g., 2.8-fold or less, 2.5-fold or less, 2.0-fold or less, 1.8-fold or less, 1.6-fold or less, 1.4-fold or less, 1.2-fold or less, or 1.1-fold or less) after 6 months of storage at 40° C. In some embodiments, the total amount of FFAs in a liquid composition formulated with PS20 described herein after 6 months of storage at 40° C. is 35 nmol / ml or less (e.g., 30 nmol / ml or less, 25 nmol / ml or less, 20 nmol / ml or less, 18 nmol / ml or less, 15 nmol / ml or less, 12 nmol / ml or less, 10 nmol / ml or less, 8 nmol / ml or less, or 5 nmol / ml or less, or any range between 5 nmol / ml and 10 nmol / ml, etc., inclusive). In some embodiments, the FFA is present in an amount that is below or at the detection limit of the FFA detection assay, eg, less than or equal to 1 nmol / ml. 【0342】 In some embodiments, the liquid risankizumab compositions described herein comprise PS80, and the stability of PS80 in such liquid risankizumab compositions is increased. 【0343】 In one embodiment, the stability of PS80 can be assessed by directly measuring the amount of PS80 in a risankizumab composition after storage for a period of time (e.g., 3 months, 6 months, 9 months, 12 months, 18 months, 24 months, 30 months, 36 months, etc.) at a particular temperature (e.g., 2°C, 4°C, 5°C, 8°C, 10°C, 12°C, 25°C, 30°C, 35°C, or 40°C). The amount of PS80 can be measured using any method known in the art, for example, using HPLC-CAD, such as the HPLC-CAD described in Example 10. In some embodiments, the period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, or more months, or any range between 3 and 36 months, 12 and 24 months inclusive, etc. 【0344】 For example, in some embodiments, at least 80% (e.g., at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of PS80 is retained after 6 months of storage at 5°C. In some embodiments, at least 60% (e.g., at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of PS80 is retained after 6 months of storage at 25°C. In some embodiments, at least 60% (e.g., at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of PS80 is retained after 6 months of storage at 40°C. 【0345】 In one embodiment, the stability of PS80 can be assessed by measuring the amount of free fatty acids (FFAs), which are degradation products of PS80, in a risankizumab composition formulated with PS80 after storage at a particular temperature (e.g., 2°C, 4°C, 5°C, 8°C, 10°C, 12°C, 25°C, 30°C, 35°C, or 40°C) for a period of time (e.g., 3 months, 6 months, 9 months, 12 months, 18 months, 24 months, 30 months, 36 months, etc.). The amount of FFAs can be measured using any method known in the art, for example, using an enzymatic FFA or LC-FFA assay, e.g., the RP-HPLC-UV method described in Example 10. In some embodiments, the period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, or more months, or any range between 3 and 36 months, 12 and 24 months inclusive, etc. 【0346】 For example, in some embodiments, the total amount of FFAs in a liquid composition formulated with PS80 described herein increases by 8-fold or less (e.g., 7-fold or less, 6-fold or less, 5-fold or less, 4-fold or less, 3-fold or less, 2-fold or less, 1.5-fold or less, 1.2-fold or less, or 1.1-fold or less) or does not increase after 6 months of storage at 5°C. In some embodiments, the total amount of FFAs in a liquid composition formulated with PS80 described herein is 45 nmol / ml or less (e.g., 40 nmol / ml or less, 35 nmol / ml or less, 30 nmol / ml or less, 25 nmol / ml or less, 20 nmol / ml or less, 18 nmol / ml or less, 15 nmol / ml or less, 12 nmol / ml or less, 10 nmol / ml or less, 8 nmol / ml or less, or 5 nmol / ml or less, or any range therebetween (inclusive), such as 5 nmol / ml to 10 nmol / ml, 5 nmol / ml to 20 nmol / ml, 5 nmol / ml to 30 nmol / ml, or 5 nmol / ml to 45 nmol / ml, etc.) after 6 months of storage at 25° C. In some embodiments, the FFAs are present in an amount that is below or at the detection limit of an FFA detection assay, e.g., less than or at 1 nmol / ml. 【0347】 In some embodiments, the total amount of FFAs in a liquid composition formulated with PS80 described herein increases by 12-fold or less (e.g., 11-fold or less, 10-fold or less, 9-fold or less, 8-fold or less, 7-fold or less, 6-fold or less, 5-fold or less, 4-fold or less, 3-fold or less, 2-fold or less, 1.5-fold or less, 1.2-fold or less, or 1.1-fold or less) after 6 months of storage at 25°C. In some embodiments, the total amount of FFAs in a liquid composition formulated with PS80 described herein is 65 nmol / ml or less (e.g., 60 nmol / ml or less, 55 nmol / ml or less, 50 nmol / ml or less, 45 nmol / ml or less, 40 nmol / ml or less, 35 nmol / ml or less, 30 nmol / ml or less, 25 nmol / ml or less, 20 nmol / ml or less, 18 nmol / ml or less, 15 nmol / ml or less, 12 nmol / ml or less, 10 nmol / ml or less, 8 nmol / ml or less, or 5 nmol / ml or less, or any range therebetween (inclusive), such as 5 nmol / ml to 10 nmol / ml, 5 nmol / ml to 20 nmol / ml, 5 nmol / ml to 30 nmol / ml, or 5 nmol / ml to 65 nmol / ml, etc.) after 6 months of storage at 25°C. In some embodiments, the FFA is present in an amount that is below or at the detection limit of the FFA detection assay, eg, less than or equal to 1 nmol / ml. 【0348】 In some embodiments, the total amount of FFAs in a liquid composition formulated with PS80 described herein increases by 2.5-fold or less (e.g., 2-fold or less, 1.5-fold or less, 1.2-fold or less, or 1.1-fold or less) or does not increase after 6 months of storage at 40° C. In some embodiments, the total amount of FFAs in a liquid composition formulated with PS80 described herein increases by 15 nmol / ml or less (e.g., 12 nmol / ml or less, 10 nmol / ml or less, 8 nmol / ml or less, 5 nmol / ml or less, or 3 nmol / ml or less, or any range between 3 nmol / ml and 5 nmol / ml, 3 nmol / ml and 10 nmol / ml, 5 nmol / ml and 10 nmol / ml, or 3 nmol / ml and 15 nmol / ml, etc., inclusive) after 6 months of storage at 40° C. In some embodiments, the FFAs are present in an amount below or at the detection limit of an FFA detection assay. 【0349】 In some embodiments, the risankizumab compositions described herein comprise poloxamer 188. In some embodiments, the P188-containing risankizumab compositions do not comprise PS20 and / or PS80. In some embodiments, the stability of P188 in the risankizumab compositions is increased relative to the stability of PS20 or PS80. 【0350】 In some embodiments, the stability of P188 can be assessed by directly measuring the amount of P188 in a risankizumab composition after storage for a period of time (e.g., 3 months, 6 months, 9 months, 12 months, 18 months, 24 months, 30 months, 36 months, etc.) at a particular temperature (e.g., 2°C, 4°C, 5°C, 8°C, 10°C, 12°C, 25°C, 30°C, 35°C, or 40°C). The amount of P188 can be measured using any method known in the art, for example, using a Pluronic F-68 colorimetric assay, such as the Pluronic F-68 colorimetric assay described in Example 17. In some embodiments, the period is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36 or more months, or any range between 3 and 36 months, 12 and 24 months, 3 and 6 months inclusive, etc. 【0351】 For example, in some embodiments, at least 85% (e.g., at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of P188 is retained after 3 months of storage at 5°C. In some embodiments, at least 65% (e.g., at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of P188 is retained after 3 months of storage at 25°C. In some embodiments, at least 60% (e.g., at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of P188 is retained after 3 months of storage at 40°C. 【0352】 For example, in some embodiments, at least 80% (e.g., at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of P188 is retained after 6 months of storage at 5°C. In some embodiments, at least 60% (e.g., at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of P188 is retained after 6 months of storage at 25°C. In some embodiments, at least 60% (e.g., at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or about 100%) of the initial amount of P188 is retained after 6 months of storage at 40°C. [Example] 【0353】 Example 1: Particles were observed upon dilution of risankizumab drug products DP1 and DP2. Risankizumab Drug Product 1 (DP1) at 90 mg / ml was developed using the process and formulation described in International Application No. PCT / US2013 / 038109. Risankizumab Drug Product 2 (DP2) at a concentration of 150 mg / ml was subsequently developed and approved by the FDA. The formulations of DP1 and DP2 are shown in Table 6 below. 【0354】 [Table 6] 【0355】 Risankizumab formulations DP1 and DP2 contain highly purified risankizumab API and are stable. However, as shown in Table 7 below, particles containing free fatty acids (FFA) were unexpectedly observed in DP1 and DP2, particularly when DP2 was diluted (e.g., 51 mg / ml and 60 mg / ml) and used to investigate the feasibility of on-body device presentation and placed under specific storage conditions. 【0356】 [Table 7] 【0357】 Example 2: Candidate hitchhiker proteins were identified by proteomic analysis. We hypothesized that particle formation in risankizumab products is due to degradation of the surfactant polysorbate 20 (PS20), which can lead to antibody aggregation. PS20 degradation was hypothesized to result from residual CHO cell esterases co-purified with the risankizumab drug substance. To identify putative hitchhiker proteins (HPs) that may have led to degradation of polysorbate-20 in the risankizumab drug product, host cell proteins (HCPs) from DP1 and DP2 bulk drug substance (BDS) samples were enriched and subsequently analyzed by LC-MS / MS. 【0358】 Multiple risankizumab BDS samples from each DS process were pooled to generate representative BDS material for HCP enrichment and identification. In total, six BDS batches from Process 1 were pooled to generate a representative DP1 BDS sample, and four DP2 BDS batches were combined to generate a DP2 BDS sample for HCP enrichment and identification testing. 【0359】 The protein concentration of each representative risankizumab BDS sample was measured and is shown in Table 8. 【0360】 [Table 8] 【0361】 method 【0362】 Immunoaffinity purification for HCP enrichment from risankizumab 【0363】 1. Coupling of anti-HCP antibodies to Sepharose beads 【0364】 a. Preparation of buffer solutions: 【0365】 Acidification buffer: 1 mM HCl, 【0366】 Coupling buffer: 200 mM NaHCO3, 500 mM NaCl, pH 9.0, 【0367】 Blocking buffer: 1 M ethanolamine, pH 8.0, 【0368】 Wash buffer: 100 mM sodium acetate, 500 mM NaCl, pH 4.0. 【0369】 b. Buffer exchange of anti-HCP antibody 【0370】 Anti-total HCP antibody (CRO: Biogenes®): 1.99 mg / mL, 【0371】 Anti-LMW HCP antibody (CRO: Covance (registered trademark)): 5.19 mg / ml, 【0372】 Dialysis machine: Thermo Scientific Pro #66810. 【0373】 3 mL of anti-total HCP antibody and 3 mL of anti-LMW HCP antibody were each buffer exchanged in 4 L of coupling buffer (described above) by dialysis overnight in a cold room using slow mixing (approximately 60 rpm). 【0374】 c. Conditioning of cyanogen bromide (CNBR)-activated Sepharose beads (Cytiva catalog number 71-5000-15 AF) 【0375】 0.5 g of CNBR Sepharose beads were weighed and placed in a polyprep chromatography column (Bio-Rad catalog number 731-1550). The Sepharose beads were suspended in 5 mL of 1 mM ice-cold HCl. The column was inverted to ensure the Sepharose beads were completely wetted (approximately 10 minutes). The column was placed in a 15 mL centrifuge tube and centrifuged at 200 g for 7 minutes to dry the beads (Beckman Avanti J-15R). The CNBR Sepharose beads were washed three more times with 1 mM ice-cold HCl. 【0376】 d. Anti-HCP antibody coupling and washing 【0377】 Four milliliters of buffer-exchanged anti-total HCP antibody and anti-LMW HCP antibody were added to two separate polypreparative chromatography columns containing the conditioned CNBR-activated Sepharose beads obtained in step c. To allow binding of the anti-LMW or anti-total HCP antibody to the CNBR-activated Sepharose beads, the two columns were kept in a cold room overnight with slow mixing on a rocker. The polypreparative chromatography columns were centrifuged, and unbound anti-HCP antibody was removed from the Sepharose beads by centrifugation at 200 g (Beckman Avanti J-15R) for 7 minutes. 【0378】 Five milliliters of blocking buffer was added to each column and mixed overnight in a cold room at low speed on a bench rocker to block the antibody-immobilized Sepharose beads. The blocking buffer was removed by centrifugation (as described in step c). The columns were washed alternately with coupling buffer and wash buffer (four times each, 5 mL per wash). Finally, the columns were conditioned in 1x PBS buffer by adding 5 mL of 1x PBS twice and eluting. The Sepharose beads (antibody-bound) were suspended in PBS buffer and stored in the columns at 4°C until use. 【0379】 2. Packing of immunoaffinity columns for HCP purification / enrichment 【0380】 The suspension Sepharose beads (antibody-bound) were transferred from the polyprep chromatography column to a Tricorn 5 / 50 column (Cytiva product code: 28406409; approximately 1 mL column volume (CV)). Agitation was minimized to prevent air bubbles from being introduced into the Sepharose bead packing within the Tricorn column. The Tricorn column was connected to a peristaltic pump P1 (Cytiva) to set up an immunoaffinity purification system. The Tricorn column was conditioned by driving 20 mL (20 CV) of PBS buffer using the P1 pump to drive the liquid flow at a flow rate of 0.5 mL / min. The Tricorn column packed with Sepharose beads was stored at 4°C. 【0381】 3. HCP Coupling, Washing, and Elution from the Purification Column 【0382】 Pooled risankizumab BDS samples (DP1 and DP2) were purified sequentially using two Tricorn immunoaffinity columns (anti-total HCP and anti-LMW HCP columns) loaded as described above, shared with two separate P1 pumps. For each risankizumab BDS sample, the following procedures and materials were used for HCP coupling, washing, and elution from the immunoaffinity columns. 【0383】 a. Preparation of buffer solutions 【0384】 Protein binding buffer: 1× PBS (pH 7.4); 【0385】 Washing buffer: 1x PBS, 0.05% Tween 20 (pH 7.4); 【0386】 Elution buffer: 100 mM glycine, 400 mM arginine (HCl) (pH 2.7); 【0387】 Neutralization buffer: 1 M Tris-HCl (pH 8.5). 【0388】 A total of 4.5 g of risankizumab BDS sample was prepared (50 mL at 90 mg / mL or 30 mL at 150 mg / mL, depending on the risankizumab concentration in the pooled BDS sample) and purified on each Tricorn immunoaffinity column (two columns total). 50 (30) mL of the pooled BDS sample from DP1 or DP2 was aliquoted into five aliquots (10 mL or 6 mL each) for each loading cycle of the risankizumab BDS sample and washed on the immunoaffinity column (five cycles total, each aliquot for one cycle). For each cycle, 10 mL (or 6 mL) of the risankizumab BDS sample was circulated through the Tricorn immunoaffinity column for 40 minutes at a flow rate of approximately 0.5 mL / min. The Tricorn column was washed with approximately 20 CV of wash buffer delivered by the P1 pump. The cycling and washing steps were repeated for the other four aliquots of risankizumab BDS sample in each Tricorn immunoaffinity column. After five cycles of loading and washing the risankizumab BDS sample on each Tricorn column, each column was washed with an additional 10 CV of wash buffer. Bound proteins (HCP and risankizumab molecules) were eluted from each Tricorn column at the same flow rate of approximately 0.5 mL / min using 15 mL (CV) of elution buffer delivered by the P1 pump. The eluate was neutralized to approximately pH 7 (measured with a pH probe) with neutralization buffer (1 mL of eluate mixed with 200 μL of neutralization buffer). Neutralization should be performed after the purified eluate leaves the column. Protein concentration was measured in each neutralized eluate from the Tricorn immunoaffinity column for each risankizumab BDS sample using a Lunatic spectrophotometer. Concentrations should be below the detection limit. 【0389】 4. Concentration and Purification of Eluate from Immunoaffinity Column 【0390】 The purified eluate (approximately 15 mL) was concentrated to a total volume of approximately 0.5 mL using a 15 mL 3K MWCO (Millipore, catalog number UFC900324). The 0.5 mL eluate was further concentrated to a final volume of approximately 100 μL using a 0.5 mL, 3K MWCO (Millipore, catalog number UFC500324). The total protein concentration in each 100 μL concentrated eluate was measured using a Lunatic spectrophotometer. The total protein concentration should be approximately 1 mg / mL. Similar purification and concentration procedures were performed for all four representative risankizumab BDS samples using the same immunoaffinity purification columns (anti-LMW and anti-total HCP columns). A total of eight elution samples were collected from the purification and concentration of all four pooled risankizumab BDS samples for LC-MS / MS analysis. 【0391】 LC-MS / MS method for HP identification 【0392】 1. Sample Denaturation and Reduction 【0393】 60 μL of 8 M urea and 8.8 μL of 1 M DTT were added to each 20 μL eluate sample, and each sample was incubated at 37° C. for 30 minutes with shaking at 450 rpm. 【0394】 2. Sample Alkylation and Digestion 【0395】 200 μL of 8M urea was added to a 0.5mL 30kDa MWCO centrifugal filter (Millipore, catalog number MRCF0R030). The reduced sample was then added to the filter and centrifuged at 14,000×g for 15 minutes. 400 μL of 8M urea was added to the filter and centrifuged at 14,000×g for 15 minutes. The flow-through from the collection tube was discarded. 100 μL of 50mM iodoacetamide solution was added, mixed in a thermomixer at 600 rpm for 1 minute, and incubated at room temperature for 20 minutes without mixing. The filter was centrifuged at 14,000×g for 10 minutes. 100 μL of 8M urea was added to the filter and centrifuged at 14,000×g for 15 minutes. This step was repeated once. 100 μL of 50 mM ammonium bicarbonate was added to the filter unit and centrifuged at 14,000 × g for 10 minutes. This step was repeated once. The filter was transferred to a new collection tube. 【0396】 35 μL of 50 mM ammonium bicarbonate was added, followed by 1 μL of 0.4 μg / μL trypsin (Thermo Scientific, catalog number 90057, enzyme-to-protein ratio 1:50) and mixed in a thermomixer at 600 rpm for 1 minute. The filters were incubated overnight at 37°C in the thermomix. The tubes were wrapped in parafilm to prevent evaporation. The next day, the filters were centrifuged at 14,000 × g for 10 minutes. 40 μL of 50 mM ammonium bicarbonate was added, and the filters were centrifuged at 14,000 × g for 10 minutes. The samples were acidified with formic acid to ensure a pH < 1. Peptide concentrations were measured by Bradford colorimetric assay (Thermo Scientific, catalog number 23250). 【0397】 3.LC-MS / MS analysis 【0398】 For each digested sample, 800 ng was injected onto a Bruker nanoEcute™ ultra-high pressure liquid chromatography (UHPLC) system equipped with an Aurora Series UHPLC C18 column, 120 Å, 25 cm × 75 μm ID, 1.6 μm (Ionoptics, catalog number AUR2-25075C18A-CSI) maintained at 50° C. Peptides were eluted into the mass spectrometer using a gradient of mobile phase A (0.1% formic acid in water) and mobile phase B (0.1% formic acid in acetonitrile) shown in Table 9 at a flow rate of 250 nL / min. 【0399】 [Table 9] 【0400】 Data were acquired using a Burker timsTOF Pro QTOF mass spectrometer operated in positive ion mode, scanning from 100 to 1700 m / z in PASEF mode. The ion mobility resolution was set to 0.60–1.35 V·s / cm² over a 100 ms ramp time. For each cycle, 10 PASEF MS / MS scans were performed at 14,500 intensity units toward the precursor to reach a near 100% duty cycle. A polygon filter was applied to m / z and ion mobility to exclude low m / z singly charged ions from MS / MS fragmentation. Collision energy was ramped stepwise according to ion mobility. Data were searched against the CHO proteome database using MSFragger V17.1. 【0401】 result 【0402】 [Table 10] 【0403】 Affinity purification identified various hitchhiker proteins in pooled risankizumab BDS samples, including putative phospholipase B-like 2 (PLBL2), acid ceramidase, isoamyl acetate-hydrolyzing esterase 1, sphingomyelin phosphodiesterase, hepatic carboxylesterase-like isoform 1 (CES1), hepatic carboxylesterase 4, ester hydrolase C11orf54 homolog isoform 1, sialic acid O-acetylesterase-like (SIAE), calcineurin-like phosphoesterase domain-containing protein 1, and peroxiredoxin-6-like (Prdx6) (Figure 1 and Table 10). Some of these proteins, such as PLBL2 and acid ceramidase, have previously been reported to be present in other antibody drug substances (Graf et al. (2021) J. pharm. Sci. 110:3358-3567). 【0404】 Example 3: Development of Risankizumab Drug Product with Improved Stability The present inventors hypothesized that the presence of certain types of hitchhiker proteins (e.g., host cell lipases) derived from host cells causes particle formation in diluted risankizumab drug products. To solve this problem and improve the stability and shelf life of risankizumab products, as well as to further improve their quality, a novel risankizumab drug substance, Process 4 DS, was developed. An exemplary purification process for producing Process 4 DS is described below. 【0405】 CHO cells expressing risankizumab were thawed and cultured to increasing volumes in shake flasks, cell bags, and seed bioreactors to provide sufficient cells to inoculate the production bioreactor. The cell culture broth was harvested by centrifugation and filtration to efficiently remove cells and provide a clarified harvest for further product purification. The clarified harvest was then processed through a series of chromatography steps: viral inactivation, viral filtration, concentration and buffer exchange by tangential flow filtration, and final formulation. The purification process was developed to reduce host cell lipases by screening various reagents and conditions, including, but not limited to, Protein A chromatography wash schemes and wash buffers, depth filters, chromatography column resins (e.g., AEX resin, CEX resin, MM resin, and / or HIC resin), and / or ultrafiltration and dialysis (UF / DF) process conditions. Reagents and conditions that effectively reduce specific host cell lipase levels in purified risankizumab drug substance (DS) with acceptable yield trade-offs, as measured by ELISA, were adapted to establish two optimized purification processes, referred to herein as Process 3 and Process 4. In addition to the Process 4 purification process, the upstream cell culture process was further modified to enhance culture longevity, productivity, and robustness. 【0406】 A general overview of the purification process for Process 4 is shown in Figures 14A and 14B. Specifically, cell culture broth was harvested and clarified by centrifugation and depth filtration using an X0HC depth filter. The clarified harvest was first purified by affinity chromatography using MabSelect SuRe™ Protein A resin. The eluate was subjected to low-pH inactivation using phosphoric acid and then depth filtration using X0HC and D0HC depth filters. The risankizumab antibody sample was then purified by Capto™ Adhere mixed-mode chromatography. The flow-through was further purified by cation exchange chromatography using Poros™ XS resin. The eluate was subjected to viral filtration. Ultrafiltration / dialysis (UF / DF) was then performed by directly spiking the load with a high-salt solution, followed by spiking 8 DV without salt. The purified bulk drug substance (BDS) was then formulated and appropriately stored. 【0407】 The detailed parameters of the different steps of Process 4 are listed below. 【0408】 Primary collection 【0409】 Primary recovery by centrifugation and depth filtration was used to remove cells and cell debris from the production bioreactor tank. The 3000 L production bioreactor served as a feed tank to a 710 Alpha Laval centrifuge. Centrifugation was performed at a setpoint of 5555 rpm, with a feed rate of 30 L / min and discharge intervals of 215 seconds for GMP1-GMP3 and 252 seconds for GMP4, respectively. The pellet was then separated into eighteen 1.1 m 2 A filter train consisting of 1 Millipore X0HC Media Pod unit followed by two Sartopore2 30-inch filters (1.8 m each) 2 The solution was passed through a 0.45 μm / 0.2 μm capsule filter. 【0410】 The harvest from the bioreactor was centrifuged and depth filtered, after which the filter was flushed with a target weight of 396 kg of 50 mM sodium acetate, pH 5.5 buffer. Centrifugation and filtration of the harvest were performed as a single unit operation. Filtration was performed at ambient temperature (18-25°C) in the fermentation suite. Prior to filtration, the harvest temperature was cooled to 18-22°C with a setpoint of 20°C. The filtrate was collected in a 3000 L recovery tank, cooled to 2-8°C, and allowed to remain for up to 3 days. 【0411】 MabSelect SuRe Protein A Chromatography 【0412】 MabSelect SuRe Protein A chromatography was used to capture risankizumab Process 4 DS from the clarified harvest and reduce the amount of process-related impurities. The MabSelect SuRe self-packed column (GE Healthcare) was 60 cm in diameter with a target volume of approximately 62.0 L (21-23 cm bed height). Operations were carried out in the fermentation suite at ambient temperature (18-25°C) using the process parameters shown in Table 11A below. 【0413】 The MabSelect SuRe column was operated in bind-and-elute mode. Three cycles of MabSelect SuRe chromatography were required to process each batch. The column was equilibrated with 50 mM sodium acetate, pH 5.5, and then loaded with 13–35 g of risankizumab per L of resin. After loading, three wash steps were performed. Wash 1 was 50 mM sodium acetate, pH 5.5. Wash 2 was 50 mM Tris, 1 M arginine, pH 8.0, and Wash 3 was 50 mM sodium acetate, pH 5.5. Elution was performed with 50 mM sodium acetate, pH 3.5. The column was then regenerated with 0.2 M sodium hydroxide and re-equilibrated with equilibration buffer before loading the next cycle. 【0414】 The load material (2-8°C) was not warmed before loading onto the column. Collection of the eluate peak started at 0.2 OD and descended to 0.2 OD (wavelength 280 nm, path length 1 mm). The eluate was placed into a 300 L portable stainless steel tank or a 500 L disposable mixing (SUM) system, then passed offline through a 0.45 / 0.2 μm filter into the collection vessel. One Sartopore2 30-inch (1.8 m each) filter was used. 2 ) 0.45 μm / 0.2 μm capsule filters were used for eluate filtration. Each eluate filter was used for three daily cycles of MabSelect SuRe chromatography. MabSelect SuRe eluate was collected in a 700 L portable stainless steel tank or a 1000 L single-use mixing (SUM) system and allowed to stand at 9-25°C for up to 1 day or cooled to 2-8°C for up to 3 days before proceeding to low-pH inactivation. 【0415】 [Table 11] 【0416】 pH inactivation and POD filtration 【0417】 The purpose of the pH inactivation step was to inactivate any adventitious viruses that may be present. The pH inactivation step was carried out in the fermentation suite at ambient temperature (18-25°C). 【0418】 The pH of the Protein A eluate was adjusted to 3.5 ± 0.1 (measured at 18–25 °C) using 0.5 M phosphoric acid. After a 60–90 min hold period, the inactivated material was neutralized to pH 8.0 ± 0.1 (measured at 18–25 °C) using 2.0 M Tris. The conductivity of the material should be within the range of 3.8–4.8 mS / cm (measured at 24–26 °C) for subsequent filtration; therefore, no dilution was required prior to POD filtration. 【0419】 Depth filtration was used to remove particles and reduce impurity levels in the process stream. The filter trains consisted of two 1.1 m 2 Millipore D0HC Media Pod unit and five 1.1m 2 Millipore X0HC Media Pod Unit and Sartopore 2 30-inch (1.8 m each) 2 The filter train consisted of a 0.45 μm / 0.2 μm capsule filter. To increase step yield, the number of DOHC and XOHC filters was reduced to one and four, respectively, in GMP3 and GMP4. The filter train was set to approximately 37.5 L / m 2 The contents of the feed tank were then equilibrated with approximately 20.0 L / m of 25 mM Tris, 25 mM sodium chloride, pH 8.0, and then filtered. 2 The column was rinsed with 25 mM Tris, 25 mM sodium chloride, pH 8.0. The filtrate was collected in a 700 L portable stainless steel tank or a 1000 L SUM system and advanced to Capto Adhere chromatography the same day. 【0420】 Capto Adhere Chromatography 【0421】 A Capto Adhere chromatography step was used to reduce the impurity levels in the process stream. The column packed with Capto Adhere resin was 45 cm in diameter with a target volume of 19.1 L (12 cm bed height). Operation was carried out in the purification suite at ambient temperature (18-25°C) using the process parameters shown in Table 11B below. 【0422】 The Capto Adhere column was operated in flow-through mode. One cycle of Capto Adhere chromatography was required to process each batch. The column was first pre-equilibrated with 2 M sodium chloride and then equilibrated with 25 mM Tris, 25 mM sodium chloride, pH 8.0. Risankizumab (150–300 g / L resin) was loaded onto the column and then washed with 260 mM Tris, pH 8.0. The column was regenerated with 0.1 M acetic acid, pH 2.9, and 2 M sodium chloride. The column was sterilized with 1 M sodium hydroxide after each batch and stored in 0.1 M sodium hydroxide. 【0423】 The load material was maintained at 18-25°C before loading onto the column. During loading, flow-through product was collected from 1 OD on the peak front and ended at 5 OD on the peak tail during washing (wavelength 280 nm, path length 1 mm). Capto Adhere flow-through was collected in a 1000 L portable stainless steel tank or a 1000 L SUM system. 【0424】 Capto Adhere FTW was adjusted to a target pH of 5.25 on the day of Capto Adhere chromatography. To prepare the Poros XS load, the Capto Adhere FTW material was titrated to pH 5.25 + 0.1 (measured at 18-25°C) using 2 M acetic acid, and the conductivity was adjusted to 4.5-7.5 mS / cm with WFI as needed. The adjusted Capto Adhere FTW was then loaded onto a Sartopore2 30 in (1.8 m) column. 2 The filtered Capto Adhere FTW was filtered through a single 0.45 μm / 0.2 μm capsule filter. The filtered, prepared Capto Adhere FTW could be kept at 9–25°C for up to 1 day or chilled to 2–8°C for up to 3 days before proceeding to Poros XS chromatography. 【0425】 [Table 12] 【0426】 Poros XS Chromatography 【0427】 A Poros XS chromatography step was used to reduce basic species and treat related impurities such as host cell proteins and spillover eluted Protein A. The column packed with Poros XS resin had a diameter of 60 cm and a target volume of 56.5 L (bed height of 20 cm). The operation was carried out in the purification suite at ambient temperature (18-25°C) using the process parameters shown in Table 11C below. 【0428】 The Poros XS column was operated in both binding and elution modes. Two cycles of Poros XS chromatography were required to process each batch. The column was equilibrated with 50 mM sodium acetate, 31 mM sodium chloride, pH 5.25. The column load range was 25–50 g risankizumab per L of resin. The wash step was 50 mM sodium acetate, 31 mM sodium chloride, pH 5.25. Elution was performed with 50 mM sodium acetate, 181 mM sodium chloride, pH 5.25. The pH of the elution buffer for GMP1–GMP3 was close to target, but to improve the Poros XS step yield, the pH of the elution buffer for GMP4 was adjusted to 5.34 using 5 M sodium hydroxide, which is at the higher end of the elution pH batch's recorded range (5.15–5.35). Before the next cycle, the column was regenerated with 25 mM Tris, 3 M sodium chloride, pH 8.5. Finally, the column was sterilized with 1.0 M sodium hydroxide and stored in 0.1 M sodium hydroxide. At the end of the last cycle of each batch run, the column was sterilized and stored. 【0429】 The Poros XS load was measured on the day of Poros XS chromatography using Sartopore2 30-inch (1.8 m each) columns. 2The eluate was filtered through one 0.45 μm / 0.2 μm capsule filter. Collection of the eluate peak began at 1 OD from the peak front and ended at 5 OD from the peak tail during elution (wavelength 280 nm, path length 1 mm). Two Sartopore 2 30-inch (1.8 m each) filters were used for eluate filtration. 2 A 0.45 μm / 0.2 μm capsule filter was used to pass the eluate through the 0.45 / 0.2 μm filter as it left the chromatography skid and into the collection vessel. This filter was used for two Poros XS chromatography cycles on the same process day. The Poros XS eluate was collected in a 500 L portable stainless steel tank or a 1000 L SUM system and held at 2–25°C for up to 5 days before proceeding to nanofiltration. 【0430】 [Table 13] 【0431】 Virus filtration 【0432】 Viral filtration provided the ability to remove adventitious viruses greater than 20 nm in size. This process was carried out at ambient temperature (18-25°C) in the purification suite. 【0433】 The virus filtration filter train consisted of two parallel Millipore Virosolve Pro Magnus 2.1 shields (0.51 m each). 2 ) 0.1 μm capsule filter, in parallel with two Millipore Virosolve Pro Mgnus 2.1 filters (each 0.51 μm 2 ), and a single Sartopore2 30-inch (1.8m) 2The filter consisted of a 0.45 μm / 0.2 μm capsule filter. Prior to product filtration, each prefilter and Virosolve nanofilter was flushed with at least 102 L of WFI, followed by at least 26 L of 50 mM sodium acetate, 181 mM sodium chloride, pH 5.25. Product filtration was performed using a Quatroflow pump with a target nanofilter pressure of 23 psig and an upper limit of 32 psig. The post-filtration flush was 20 L of 50 mM sodium acetate, 181 mM sodium chloride, pH 5.25. The viral filtrate may be held at 2-25°C for up to 5 days. 【0434】 UF / DF 【0435】 A UF / DF step was used to concentrate the product and dialyze it into the final formulation buffer. The process was carried out in the purification suite at ambient temperature (18-25°C) utilizing a 30 kD Millipore Pellicon3 Biomax UF module, D screen, using the process parameters shown in Table 11D below. 【0436】 The load material was diluted 10x with 5 M sodium chloride (9 parts nanofiltrate to 1 part 5 M sodium chloride), and the pH of the load material was then adjusted to 5.45 ± 0.1 with 2 M sodium acetate (measured at 18-25 °C). The conditioned load material was filtered through a single Sartopore2 30-inch (1.8 m) UF / DF membrane before being loaded onto the membrane. 2 ) Filtered through a 0.45 μm / 0.2 μm capsule filter into a recirculation tank. 【0437】 UF / DF is Skid Z-2300, 1.14m 2 8 membranes, total 9.12m 2The UF / DF load was concentrated to a target of 50 g / L, then dialyzed against 0.002% (w / v) sodium chloride, followed by concentration to 235 g / L. The retentate was removed from the ultrafiltration membrane and system and passed through a single Sartopore2 10-inch 0.45 / 0.2 μm (0.45 m) filter as it entered a collection vessel, a 100 L impulse mixer system. 2 The ultrafiltration system was rinsed with approximately 5 kg of 0.002% sodium chloride to recover any product retained in the system. Both rinse 1 and rinse 2 were passed through the same Sartopore 2 10-inch 0.45 / 0.2 μm (0.45 m) sterile filter. 2 ) through a sterile filter and transferred separately to Rinse 1 and Rinse 2 collection bags. After collection of the rinses, the retentate was diluted with the appropriate amounts of Rinse 1 and Rinse 2 to achieve a concentration target of 200 g / L. 【0438】 The retentate pool was formulated by adding 5x formulation buffer, 50 mM acetate, 925 mM trehalose, 0.1% Tween 20, pH 5.70. The final bulk drug substance was diluted to 150 g / L risakizumab with 1x formulation buffer, 10 mM acetic acid, 185 mM trehalose, 0.02% Tween 20, pH 5.70. The formulated UF / DF retentate was then filtered through a 0.22 μm Millipak 200 sterile filter (0.1 m 2 The final formulated UF / DF retentate can be held at 9-25°C for up to 1 day and cooled at 2-8°C for up to 5 days before proceeding to the final bagging step. 【0439】 [Table 14] 【0440】 bagging 【0441】 The purpose of the bagging was to package and store the final bulk drug substance. The operation was performed in the purification suite at ambient temperature (18-25°C). The filtered formulated UF / DF retentate was pumped into sterile 6 L Celsius FFT bags. The bags were filled to a volume of approximately 6 kg. 【0442】 The parameter targets for Process 4 are summarized below in Table 11E. 【0443】 [Table 15-1] [Table 15-2] [Table 15-3] 【0444】 Risankizumab produced by Process 3 and Process 4 was formulated to produce Drug Product 3 (DP3) and Drug Product 4 (DP4), respectively, according to Table 12 shown below. 【0445】 [Table 16] 【0446】 In a 60 mg / ml presentation of DP3, no visible or shiny particles were observed over 24 months of stability at 4°C. 【0447】 The 51 mg / ml presentation further confirmed the improved stability of DP3 compared to DP2. 【0448】 Shiny particles were observed at 3 months with 51 mg / mL DP2 (Table 7), whereas no visible product-related particles were observed at 6 months with 51 mg / mL DP3. 【0449】 Similarly, no visible or shiny particles were observed in the 60 mg / ml DP4 vial presentation over 18 months of stability at 4°C. 【0450】 Example 4: Identification of PLA2 by LC-MS proteomics Several types of phospholipases, including phospholipase A2 group XV (PLA2 G15), have been identified as potential factors contributing to the degradation of polysorbate 20. These lipases can be present at very low abundance in DSs, complicating their detection by LC-MS / MS. Enriching lipase levels through immunoaffinity purification using immobilized antibodies against specific lipases can increase the abundance levels required for detection by LC-MS / MS. In this study, immunoaffinity purification was used to enrich PLA2 in DP1, DP2, DP3, and DP4 DSs. The enriched DSs were analyzed by enzyme-linked immunoassay (ELISA) and LC-MS / MS. 【0451】 Multiple batches of DS were pooled to generate material for PLA2 enrichment. The protein concentration of each DS pool is shown in Table 13. 【0452】 [Table 17] 【0453】 1. Immunoaffinity purification (chromatography) of PLA2 【0454】 A. Buffer exchange of PLA2 antibody 【0455】 (a) Buffer: 【0456】 (1) Acidification buffer: 1 mM HCl; 【0457】 (2) Coupling buffer: 200 mM NaHCO3; 500 mM NaCl, pH = 9.0; 【0458】 (3) Blocking buffer: 1 M ethanolamine; pH = 8.0; 【0459】 (4) Washing buffer: 100 mM sodium acetate; 500 mM NaCl, pH = 4.0; 【0460】 (5) PLA2 antibody: 1mg / mL. 【0461】 (b) Buffer exchange for PLA2 antibody. 【0462】 3.5 μg of PLA2 antibody was thawed at room temperature. The PLA2 antibody buffer was exchanged for coupling buffer using a Slide-A-Lyzer™ Dialysis Cassette 10K MWCO (Thermo Fisher Scientific, Cat. No. 66810). The dialysis cassette was suspended in 4 L of coupling buffer overnight at 4°C and 60 rpm. The buffer-exchanged PLA2 antibody was collected in a clean microfuge tube and proceeded to Section B below. 【0463】 B. Conditioning of cyanogen bromide (CNBR)-activated Sepharose beads 【0464】 Weighed 0.5 g of CNBR-activated Sepharose beads (Cytiva, catalog number 71-5000-15 AF) and packed the beads into a polyprep chromatography column (Bio-Rad, catalog number 731-1550). The beads were suspended in 5 mL of ice-cold 1 mM HCl solution. The column was inverted to ensure the beads were fully hydrated (approximately 10 minutes). The column was placed in a 15 mL conical tube and centrifuged at 200 g (Beckman Avanti J-15R) for 7 minutes to dry the beads. The previous two steps were repeated, and the beads were washed three more times with ice-cold 1 mM HCl solution. After the washing steps, the beads were kept dry. 【0465】 C. PLA2 antibody coupling 【0466】 The concentration of the buffer-exchanged PLA2 antibody from section A was determined using a Lunatic UV / Vis multicolor spectrophotometer with an extinction coefficient of 1.0 M-1 cm-1 (E1% = 10). The concentration of PLA2 antibody before coupling to CNBR Sepharose beads was 0.78 mg / mL. 4 mL of PLA2 antibody was added to the column packed with CNBR Sepharose beads. If necessary, the PLA2 antibody volume was increased to 4 mL using coupling buffer. The column was placed on a bench rocker and rocked at low speed overnight at 4°C to complete the antibody-bead coupling. The column was centrifuged at 200 g for 7 minutes to remove unbound antibody. The concentration of PLA2 antibody remaining in the column filtrate was determined. The concentration of PLA2 antibody remaining in the filtrate was 0.01 mg / mL. 5 mL of blocking buffer was added to the column, and the column was rocked at low speed overnight at 4°C. The column was centrifuged at 200g for 7 minutes to remove the blocking buffer. 5mL of coupling buffer was added to the column, and the column was centrifuged at 200g for 7 minutes to remove the buffer; this step was repeated three more times. 5mL of wash buffer I was added to the column, and the column was centrifuged at 200g for 7 minutes to remove the buffer. This step was repeated three more times for further washing. 5mL of PBS buffer was added to condition the column, and the column was centrifuged at 200g for 7 minutes to remove the buffer; this step was repeated, keeping the beads suspended in 5mL of PBS buffer. If not proceeding to the next step, the column was kept at 4°C. 【0467】 D. Packing of immunoaffinity columns for PLA2 enrichment 【0468】 A Tricorn 5 / 50 column (Cytiva, Cat. No. 28406409) was rinsed in 20% ethanol for at least 1 minute at room temperature. The column volume (CV) is approximately 1 mL. 【0469】 The beads (coupled with PLA2 antibody) were transferred from the polyprep chromatography column to a Tricorn 5 / 50 column. Agitation was minimized to avoid introducing air bubbles while packing the beads into the Tricorn column. The immunoaffinity column was equilibrated with 20 CV of PBS, pH 7.4, at 0.5 mL / min. This immunoaffinity column was either used in the next step described below or stored at 4°C until use. 【0470】 Elution of PLA2 from E.DS samples 【0471】 a.PLA2 enrichment 【0472】 Five 4.5g aliquots of each DS sample were prepared and loaded onto the immunoaffinity column. One aliquot was used per circulation: DP1: 10 mL per aliquot, 50 mL total; DP2: 6 mL per aliquot, 30 mL total; DP3: 6 mL per aliquot, 30 mL total; DP4: 6 mL per aliquot, 30 mL total. The column was equilibrated with 10 CV of PBS, pH 7.4. One aliquot of the DP1 BDS sample was recirculated through the immunoaffinity column at 0.5 mL / min for 40 minutes. The column was washed with 20 CV of PBS, 0.05% Tween 20, pH 7.4, at 0.5 mL / min. The recirculation and wash steps in this section were repeated for four additional aliquots of DS. The column was eluted in a 10 mL conical tube with 10 CV of 100 mM glycine, 400 mM arginine-HCl, pH 2.7, at 0.5 mL / min. The eluate was immediately neutralized with 1.5 mL of 1 M Tris-HCl (pH 8.5). The concentrated sample was kept on ice during this procedure. The neutralized eluate was confirmed to be approximately pH 7.0 using pH strips. The elution and neutralization steps were repeated for each DS pool. 【0473】 F. Concentration of concentrated samples 【0474】 Each eluate was transferred to an Amicon concentrator (3K MWCO, 15 mL; Millipore, catalog number UFC900324) and centrifuged at 4,000 g for 1 h at 4° C. Protein concentrations in the concentrated, neutralized eluates were measured by A280 using a Lunatic UV / Vis multicolor spectrophotometer (E1%=15.2). 【0475】 2.PLA2 ELISA 【0476】 PLA2 concentrations were measured by ELISA as described in other embodiments herein. 【0477】 A. Analysis of ELISA Results 【0478】 The results show that PLA2 was enriched 92-fold in DP1, 57-fold in DP2, and 80-fold in DP3. No PLA2 was detected in the DP4 sample (Table 14). 【0479】 [Table 18] 【0480】 3. LC-MS / MS method for PLA2 identification 【0481】 A. Sample Denaturation and Reduction 【0482】 8 M urea was added to 20 μg of each eluate sample to reach a urea concentration of 6 M. 1 M DTT was added to reach 10 mM. Each eluate sample was incubated at 37°C for 30 minutes with shaking at 450 rpm. 【0483】 B. Alkylation and Digestion of Samples 【0484】 200 μL of 8M urea was added to a 0.5mL 30kDa MWCO centrifugal filter (Millipore, catalog number MRCF0R030). The reduced sample was then added to the filter and centrifuged at 14,000×g for 15 minutes. 400 μL of 8M urea was added to the filter, and the filter was centrifuged at 14,000×g for 15 minutes. The flow-through was discarded from the collection tube. 100 μL of 50mM iodoacetamide solution was added, mixed in a thermomixer at 600 rpm for 1 minute, and incubated at room temperature for 20 minutes without mixing. The filter was centrifuged at 14,000×g for 10 minutes. 100 μL of 8M urea was added to the filter and centrifuged at 14,000×g for 15 minutes. This step was repeated once. 100 μL of 50 mM ammonium bicarbonate was added to the filter unit and centrifuged at 14,000 × g for 10 minutes. This step was repeated once. The filter was transferred to a new collection tube. 35 μL of 50 mM ammonium bicarbonate was added, followed by 1 μL of 0.4 μg / μL trypsin (Thermo Scientific, catalog number 90057, enzyme-to-protein ratio 1:50) and mixed in a thermomixer at 600 rpm for 1 minute. The filter was incubated in the thermomix at 37°C overnight. The tube was wrapped in parafilm to prevent evaporation. The next day, the filter was centrifuged at 14,000 × g for 10 minutes. 40 μL of 50 mM ammonium bicarbonate was added, and the filter was centrifuged at 14,000 × g for 10 minutes. The sample was acidified with formic acid to ensure a pH < 1. Peptide concentrations were measured by Bradford colorimetric assay (Thermo Scientific, Cat. No. 23250). 【0485】 C.LC-MS / MS analysis 【0486】 For each digested sample, 400 ng was injected onto a Waters AQCUITY UPLC M-Class system equipped with a Double nanoViper™ PepMap™ neo column, 2 μm, C18, 75 μm × 500 mm (Thermo Scientific, catalog number DNV75500PN) maintained at 50° C. Peptides were eluted into the mass spectrometer using a gradient of mobile phase A (0.1% formic acid / water) and mobile phase B (0.1% formic acid / acetonitrile) shown in Table 15 at a flow rate of 200 nL / min. 【0487】 [Table 19] 【0488】 Data were acquired using a Thermo Scientific Orbitrap™ Fusion™ Lumos™ mass spectrometer operated in positive ion mode. Survey scans were performed at a resolution of 240,000 from m / z 400 to 1500 with an automatic gain control (AGC) target of 4e6 and a maximum injection time of 50ms. Monoisotopic masses with 2 to 7 positive charges were selected with a minimum intensity threshold of 2.5e4 and then fragmented by higher-energy collisional dissociation (HCD). Cycle times were approximately 3 seconds. Data were searched against the CHO proteome database using Proteome Discoverer 3.0. 【0489】 As shown in Table 16, nanoLC-MS / MS analysis identified PLA2G15 (UniProt ID: G3HKV9) in the eluates of concentrated DS from DP1 and DP2 (≧2 unique peptides for protein identification). 【0490】 [Table 20] 【0491】 conclusion 【0492】 In this study, PLA2G15 protein was successfully enriched by immunoaffinity purification and then detected in DP1 and DP2 DSs using LC-MS / MS. One unique PLA2G15 peptide was detected in DP3, which was below the threshold required for protein identification, and no unique PLA2G15 peptides were detected in DP4 DSs. These findings are consistent with the degradation of PS20 observed during long-term storage or accelerated stability in DP1 and DP2 DSs, while no degradation of PS20 was observed in DP3 and DP4. 【0493】 Example 5: Identification of PLA2 by Western blot analysis Phospholipase A2 group XV (PLA2G15) levels in different risankizumab DS samples were also assessed by Western blot. 【0494】 Risankizumab BDS samples were obtained by pooling batches of risankizumab DS produced by the same risankizumab process. Risankizumab was largely depleted from the samples by ultrafiltration using an Amicon filter with a 100 kDa molecular weight cutoff. Two hundred microliters of the pooled BDS sample was added to the Amicon filter and spun until 100 microliters of filtrate was obtained. The filtrate, which contained proteins with a molecular weight less than 100 kDa and some residual risankizumab, was evaluated by Western blot analysis. 【0495】 [Table 21] 【0496】 SDS-PAGE and Western blot Risankizumab samples and CHO PLA2G15 protein were run on 4-12% SDS-PAGE. Proteins were transferred to a PVDF membrane and probed with rabbit anti-PLA2G15 antibody. Anti-rabbit antibody conjugated to horseradish peroxidase was used as the detection reagent. Development was completed with chromogenic TMB substrate. 【0497】 result 【0498】 A PLA2G15 band on Western blot with a molecular weight of 47 kDa was observed in all risankizumab BDS batches except for DP4, and was slightly less abundant in the DP3 batch (Figure 2). 【0499】 Example 6: Knockout cell line data demonstrate that PLBL2 is not a problematic hitchhiker protein. To identify the specific hitchhiker proteins that lead to PS20 degradation, we generated and characterized CHO cell lines depleted of the specific hitchhiker proteins PLBL2, PLA2, or LPL. 【0500】 Cell line development and cell culture material preparation 【0501】 CHO clones expressing risankizumab were established. The top two clones were pooled equally and used as the starting cell source for CRISPR / Cas9-mediated gene knockout (KO) experiments using a ribonucleoprotein (RNP)-based approach. In a parallel workstream, three proteins of interest were individually targeted using unique guides designed for each gene in the CHO genome: phospholipase B-like 2 protein (PLBL2; NCBI:100769512), phospholipase A2 group XV (PLA2G15; NCBI:100760699), and lipoprotein lipase (LPL; NCBI:100689191). Each KO pool was harvested after CRISPR / Cas9 RNP transfection and then single-cell cloned via limiting dilution plating. One top clone per knockout target was selected based on phenotype (growth and productivity) and next-generation sequencing (NGS) data. NGS analysis showed that the top PLBL2, LPL, and PLA2G15 knockout clones had 0%, 0.13%, and 20% wild-type sequence present in the NGS preparations, respectively. The clones were then used for antibody production. The parental cell line (called wild-type, wt) was used in parallel to generate relevant control material. Cell culture harvests were clarified by centrifugation and frozen on dry ice (for all conditions). 【0502】 Refining Development 【0503】 PLBL2 KO, PLA2G15 KO, LPL KO, and wild-type process control material was thawed and purified using Process 1 (see Example 1), and ultrafiltration / dialysis (UF / DF) followed Process 4 (see Example 3). Material from each process stream was clarified by depth filtration, followed by Protein A affinity chromatography, depth filtration, anion exchange (AEX) chromatography, cation exchange (CEX) chromatography, UF / DF, and formulation with PS20 spiked at a target risankizumab protein concentration of 150 g / L. In-process intermediates were submitted for product quality evaluation. In addition to the four preparations from the purification process (the three KOs and the wild-type control), a placebo control (negative control; 10 mM acetic acid, 185 mM trehalose, 0.02% Tween 20, pH 5.70) and a DP4 BDS control (positive control purified by Process 4, see Example 3) were also included in a PS20 stability study for up to 12 months at storage temperatures of 5°C, 25°C, and 40°C, respectively. Levels of high molecular weight (HMW) by size exclusion chromatography (SEC), charge variants by weak cation exchange chromatography (WCX), low molecular weight (LMW) by non-reducing capillary electrophoresis with sodium dodecyl sulfate (CE-SDS), free fatty acids (FFA), and PS20 were monitored at various time points during the stability study. 【0504】 Analysis and Development 【0505】 Antibody preparations from knockout samples and controls were tested for the presence of the three lipases by specific ELISA methods. PLBL2 and PLA2G15 were measured using a CHO-specific in-house method, and LPL was measured using a commercially available anti-mouse ELISA kit. PS20 stability was measured using two different methods in samples incubated for different lengths of time (several time points over a 12-month period) and at different temperatures (5°C, 25°C, and 40°C). One method measured the total amount of PS20 present in the sample. PS20 was quantified directly using an RP-HPLC-CAD method. The other method assessed PS20 degradation by measuring the amount of free fatty acids, primarily lauric acid, released upon hydrolysis. This was performed using an RP-HPLC method with UV detection. Free fatty acids (FFA) were labeled with PDAM (1-pyrenyldiazomethane) for detection prior to analysis. The signal generated is directly proportional to the amount of FFA released. The quality of the risankizumab product was also assessed for samples incubated at different time points and temperatures using SEC to measure %HMW, non-reducing CE-SDS to measure %LMW, and CEX to measure charge variants. 【0506】 result 【0507】 In-process mapping results demonstrated no impact on process performance and product quality across all conditions tested. Observations at 3 months (5°C, 25°C, and 40°C) and 6 months (storage conditions of 5°C, 25°C, and 40°C) relative to the start of the study (month 0) indicated that the absence of PLBL2 in the BDS did not improve PS20 stability when compared to the relevant process control conditions (Figures 3A-3D). These trends were observed in both the CAD and FFA assay results reported by the analytical development group (Figures 3A-3D). These data indicate that PLBL2 is unlikely to be the primary hitchhiker protein responsible for PS20 degradation in risankizumab formulations. 【0508】 [Table 22] 【0509】 Example 7: Hitchhiker protein spiking studies indicate PLA2 as a problematic hitchhiker protein. PS20 is primarily composed of polyoxyethylene sorbitan laurate. However, due to the nature of the manufacturing process, commercially available PS20 is a mixture of oligomers, including polyethylene glycol, polyethylene glycol esters, polyethoxylated isosorbides, polyethoxylated sorbitans, polysorbate monoesters, polysorbate diesters, and sorbitol polyethoxylate esters (Ayorinde et al. (2000) Rapid Commun. Mass Spectrom 14:2116-2124; Li et al. (2014) Anal. Chem. 86:5150-5157; Martos et al. (2017) J. Pharm. Sci., 106:1722-1735). Commercially available PS20 lots typically contain over 3,000 different chemical components. 【0510】 Enzymes are hitchhiker proteins (HPs) present in drug substances, such as lipases and esterases. The active site of an enzyme is composed of a combination of amino acid residues with specific structures that differ among different types of enzymes. As a result, enzymes typically have different activities and specificities for different substrates. Because PS20 is a chemical mixture, different enzymes may degrade various components of PS20 at different rates. Therefore, different enzymes may produce different PS20 degradation patterns (profiles). Therefore, degradation pattern analysis can provide useful information for identifying specific enzymes as potential underlying causes of PS20 degradation. 【0511】 PLA2G15 spiking study in DP3 material and risankizumab buffer solution 【0512】 1. Method 【0513】 The PS20-CAD subspecies method was used. This method was originally developed to qualitatively determine the subspecies composition and quantify the PS20 subspecies by comparison with a PS20 manufacturing standard. The HPLC system used in this study was an Agilent 1260 II infinity HPLC equipped with a quaternary pump, a mobile phase degassing unit, a refrigerated autosampler, a temperature-controlled column compartment, and a Thermo Scientific charged aerosol detector (CAD). Data were collected using a Waters Empower acquisition system. The method for detecting PS20 using the charged aerosol detector (CAD) is also described in Example 10. 【0514】 2. Study Design 【0515】 The study design for the PLA2G15 spiking study is shown in Table 19 below. 【0516】 [Table 23] 【0517】 Because the system takes 70 minutes to complete each injection (labeled "Injection" in Table 19), injections were repeated to measure degradation patterns (profiles) at different incubation times. All samples were kept in the autosampler at 25°C throughout the study. 【0518】 3.Results 【0519】 The degradation profile of PS20 in PLA2G15-spiked DP3 was consistent with that observed in the diluted DP2 material (Figure 4A). 【0520】 PLA2G15 spike testing at various spike levels in DP4 material 【0521】 1. Method 【0522】 The PS20-CAD variant method described above was used. 【0523】 2. Study Design 【0524】 The study design for the PLA2G15 spiking study is shown in Table 20 below. 【0525】 [Table 24] 【0526】 3.Results 【0527】 PS20 subpopulation data were collected from samples incubated at 25°C on days T0, 1, 4, and 11. All samples were maintained at 5°C in the autosampler during data collection. PS20 concentration data from T0 to 1 month (1M) were also collected. A PLA2G15 spike concentration-dependent PS20 degradation rate was observed. The PLA2G15-induced PS20 degradation pattern was very similar across all PLA2G15 spike levels (Figure 4B). 【0528】 PLBL2 spike test 【0529】 1. Background 【0530】 Phospholipase B-like 2 protein (PLBL2) was detected in the DP2 material. Early literature reported that PLBL2 could induce PS20 degradation in antibody formulations. However, very high concentrations were used in this study (Dixit, et al., Journal of Pharmaceutical Science, 2016, 105:1657-1666). A recent publication suggests that PLBL2 is unlikely to be responsible for PS20 degradation in antibody formulations due to its low activity (Zhang et al., Journal of Pharmaceutical Science, 2020, 109:2710-2718). 【0531】 2. Method 【0532】 The PS20-CAD variant method described above was used. 【0533】 3. Study Design 【0534】 The DP3 drug solution was spiked with 5 μg / mL of PLBL2 enzyme. The solution was mixed and tested. All samples were tested at 25° C. The test design was similar to the PLA2G15 spiking test in DP3 material described above. Further information regarding the test design is provided below in Table 21A. 【0535】 [Table 25] 【0536】 4.Results 【0537】 After approximately 30 hours of incubation at 25°C, no detectable degradation of PS20 was observed. Typical results are shown in Figure 5. This result confirmed that the degradation activity of PLBL2 to PS20 was low. The spiked level (5 μg / mL) was much higher than the PLBL2 concentration detected in the DP2 material by the PLBL2 ELISA method. The lack of detectable degradation of PS20 after approximately 30 hours of incubation at 25°C indicates that PLBL2 is unlikely to be the primary underlying cause of PS20 degradation in the DP2 material. 【0538】 CES spike test 【0539】 1. Background 【0540】 Carboxylesterase 1 (CES1) was detected in DP2 material. It has been reported in the literature that CES1 can induce degradation of PS20 in antibody formulations (Zhang et al. Pharmaceutical Research, 2022, 39:75-87). 【0541】 2. Method 【0542】 The PS20-CAD variant method described above was used. 【0543】 3. Study design for CES spike testing 【0544】 The DP3 drug solution was spiked with 5 μg / mL of CES1 enzyme. The solution was mixed and tested. All samples were tested at 25° C. The study design was similar to the PLA2G15 spiking study in risankizumab DP3 material. More information can be found in Table 21A. 【0545】 4.Results 【0546】 Significant PS20 degradation was observed after only a few hours of incubation at 25°C. The PS20 degradation patterns (profiles) at different incubation times were similar (although the degradation levels varied). CES1 may cause significant degradation of PS20 in risankizumab formulations. However, the PS20 degradation pattern / profile induced by CES1 in risankizumab formulations differs significantly from that observed in DP2 material. While the major PS20 peak (polyoxyethylene sorbitan monolaurate) was significantly degraded by CES1, only the PS20 subspecies region with a retention time >50 min showed little degradation. A typical result is shown in Figure 6A. A superimposition of the PS20 degradation patterns (profiles) observed in DP2 and induced by CES1 is shown in Figure 6B, which shows significant differences. Therefore, CES1 is unlikely to be the primary cause of PS20 degradation in DP2 material. 【0547】 Sialic acid O-acetylesterase spike test 【0548】 1. Background 【0549】 Sialic acid O-acetylesterase (SIAE) was detected in the DP2 material. Literature has reported that SIAE can induce PS20 degradation in antibody formulations, with moderate levels of PS20 degradation observed after 5 days of incubation at 45°C, although a high enzyme concentration (5 μg / mL) was used in this study (Zhang et al. Journal of Pharmaceutical Sciences, 2021, 110:3899-3873). The reported PS20 degradation pattern differs from that observed in the DP2 material. Literature has reported that SIAE significantly reduced the major PS20 peak (polyoxyethylene sorbitan monolaurate) (Zhang et al. Journal of Pharmaceutical Sciences, 2021, 110:3899-3873), but this peak remained stable in the DP2 material tested here. 【0550】 2. Method 【0551】 The PS20-CAD variant method described above was used. 【0552】 3. Study Design for Sialic Acid O-Acetylesterase Spiking Test 【0553】 The DP3 drug solution was spiked with 5 μg / mL of SIAE enzyme. The solutions were mixed and tested. Further information regarding the study design can be found in Table 21A. 【0554】 4.Results 【0555】 Typical results are shown in Figure 7. The results confirmed that SIAE had low decomposition activity toward PS20. At a spike level of 5 μg / mL, no detectable degradation of PS20 was observed after approximately 30 hours of incubation at 25°C. The lack of detectable degradation of PS20 after approximately 30 hours of incubation at 25°C suggests that SIAE is unlikely to be the primary cause of PS20 degradation in the DP2 material. 【0556】 Peroxiredoxin 6 (PRDX6) spiking test 【0557】 1. Background 【0558】 Peroxiredoxin 6 (PRDX6) has been detected in DP2 material using LC-MS. It has been reported in the literature that PRDX6 may have PLA2-like activity. However, the PLA2-like activity of the native protein is limited to neutral pH. This activity is further enhanced in acidic environments and at neutral pH with oxidized phospholipids (Fisher (2018) Journal of Lipid Research 59:1132-1147). 【0559】 2.Material 【0560】 Recombinant human peroxiredoxin 6 protein (PRDX6) (ab87631, AB87631-100UG, 1mg / ml, Abcam, kept at -80°C before use) 【0561】 3. Method 【0562】 The PS20-CAD variant method described above was used. 【0563】 4. Study design for PRDX6 spike testing 【0564】 Risankizumab DP3 was spiked with 5 μg / mL of PRDX6 enzyme. The solutions were mixed and tested. Further information regarding the study design is provided in Table 21B below. 【0565】 [Table 26] 【0566】 5.Results 【0567】 Typical results are shown in Figure 8. The results indicated that PRDX6 had low activity in degrading PS20. At a spike level of 5 μg / mL, no detectable degradation of PS20 was observed after approximately 30 hours of incubation at 25°C. The lack of detectable PS20 degradation after approximately 30 hours of incubation at 25°C indicates that PRDX6 is unlikely to be the primary cause of PS20 degradation in the DP2 material. 【0568】 PLA2G7 spike test 【0569】 1. Background 【0570】 Phospholipase A2 group VII (PLA2G7) has been detected in DP2 material using LC-MS. It has been reported in the literature that PLA2G7 can degrade PS20 in antibody formulation solutions (Li et al. (2021) Analytical Chemistry 93:8161-8169). 【0571】 2.Material 【0572】 Recombinant human PLA2G7 / PAF-AH / LP-PLA2 protein, (5106-PL-010; 0.44 mg / mL, Bio-techne, kept at −80°C before use) 【0573】 3. Method 【0574】 The PS20-CAD variant method described above was used. 【0575】 4. Study Design for PLA2G7 Spike Testing 【0576】 Risankizumab DP3 was spiked with 5 μg / mL of PLA2G7 enzyme. The solutions were mixed and tested. Further information regarding the study design is provided in Table 21B. 【0577】 5.Results 【0578】 Typical results are shown in Figure 9. The results confirmed that PLA2G7 caused PS20 degradation with relatively high activity. However, the PS20 degradation profile caused by PLA2G7 differed significantly from that observed in the DP2 material. The results of the spike test indicated that PLA2G7 was unlikely to be the primary cause of PS20 degradation in the DP2 material. 【0579】 summary 【0580】 The PS20 degradation profiles of DP3 and DP4 materials spiked with six enzymes were examined. These six enzymes were detected in DP2 material (four by LC-MS and two by ELISA assay). Three enzymes (PLBL2, PRDX6, and SIAE) showed very low activity against PS20 degradation, even at very high concentrations. Therefore, these three enzymes may not be the primary cause of PS20 degradation in DP2 material. The enzymes CES and PLA2G7 showed moderate activity against PS20 degradation. However, the PS20 degradation profiles caused by CES and PLA2G7 were significantly different from those observed in DP2 material. Therefore, CES and PLA2G7 are also unlikely to be the primary cause of PS20 degradation in DP2 material. The PS20 degradation profile caused by PLA2G15 closely matches the PS20 degradation profile in DP2 material. This study indicates that PLA2G15 is the major enzyme responsible for PS20 degradation in DP2 drugs. 【0581】 Example 8: PLA2G15 inhibition studies confirm PLA2 as the problematic hitchhiker protein The small molecule drug fosinopril has been reported to inhibit the activity of PLA2G15 (phospholipase A2 group XV) with an IC50 of approximately 0.18 μM (Hinkovska-Galcheva et al. (2021) J. Lipid. Res. 62:100089). Fosinopril is an angiotensin-converting enzyme (ACE) inhibitor used to treat hypertension and some chronic heart failure (Murdoch et al. (1992) Drugs 43:123-140). Furthermore, liposome-PLA2G15 co-sedimentation assays suggest that fosinopril inhibits PLA2G15 activity via interference with PLA2G15 binding to the liposome surface (Hinkovska-Galcheva et al. (2021) J. Lipid. Res. 62:100089). To further confirm that PLA2G15 is the primary cause of polysorbate 20 (PS20) degradation in DP2 material, a series of PLA2G15 inhibition studies were conducted by spiking different levels of fosinopril into DP2 material. In this study, DP2 samples were aseptically spiked with different levels of fosinopril. All samples (including unspiked samples) were incubated in the dark at room temperature for two weeks. The PS20 degradation levels of these samples were tested using the PS20 subspecies method. The PS20 degradation levels in these samples were compared with a negative control (unspiked samples kept in the dark at room temperature for two weeks) and a positive control (unspiked samples kept at -80°C before testing). Fosinopril dose-dependent protection (inhibition) of PS20 degradation in DP2 material was observed. Protection (inhibition) was observed even at spiked levels below ug / mL. This result, combined with other studies, further confirmed that PLA2G15 was the underlying cause of PS20 degradation in DP2 material. The chemical structure of fosinopril is shown below. [ka] 【0582】 material: 【0583】 DP2 material (maintained at -80°C) 【0584】 Fosinopril sodium (F13085MG) (Sigma Aldrich) 【0585】 Sample preparation: 【0586】 Fosinopril sodium was dissolved in Mill-Q water at a concentration of 0.5 mg / mL. Before use, the solution was sterile filtered through a 0.22 syringe filter. During filtration, the syringe filter was flushed with Mill-Q water and then with a 0.5 mg / mL aqueous fosinopril solution. The low-concentration aqueous fosinopril solution was aseptically diluted with Mill-Q water. More detailed sample preparation can be found in Table 22. All vials were glass, and the solution was thoroughly mixed before incubation. Samples were incubated at room temperature in the dark for two weeks before testing. 【0587】 [Table 27] 【0588】 Test Method 【0589】 All samples were qualitatively tested for PS20 levels and profiles using the PS20 HPLC-CAD variant method. The solutions were transferred to HPLC vials and diluted with Milli-Q water (1:1 dilution). All HPLC vials were mixed thoroughly and then placed in the HPLC autosampler for testing. Sample descriptions and labels can be found in Table 23. [Table 28] 【0590】 result: 【0591】 Dose-dependent protection (inhibition) of PS20 degradation by fosinopril in the DP2 material solution was observed during testing. For example, even at the lowest fosinopril spike level (0.9 μg / mL), reduced PS20 degradation was observed compared to the unspiked control sample. Figure 10 shows results for a fosinopril spike level of 0.9 μg / mL. Figure 11 shows results for a fosinopril spike level of 3.8 μg / mL. Figure 12 shows results for a fosinopril spike level of 27.8 μg / mL. 【0592】 Fosinopril could be detected at high spike levels (e.g., 39-42 min in Figure 12) because it coelutes with PS20 in the chromatogram (peak around 39.5 min retention time). Further analysis was performed to evaluate its potential impact on the observed PS20 signal. Figure 13 shows the normalized results at a very high fosinopril spike level (930 μg / mL). The fosinopril peak is located around 39.5 min. Because the fosinopril spike levels in Figures 10-12 are relatively low, the impact on retention times above 42 min can be ignored. 【0593】 summary 【0594】 Dose-dependent protection (inhibition) of PS20 degradation by fosinopril spiked into DP2 material was observed. Fosinopril demonstrated protection (inhibition) even at spike levels of 0.9 μg / mL. Because fosinopril is a potent PLA2G15 inhibitor, these results in combination with other studies further confirmed that PLA2G15 is likely the underlying cause of PS20 degradation in DP2 material. 【0595】 Example 9: Risankizumab drugs DP3 and DP4 have low hitchhiker protein levels. Residual hitchhiker protein in drug substance concentrate samples was determined by enzyme-linked immunosorbent assay (ELISA) as described below. 【0596】 PLA2 ELISA 【0597】 1. Principles 【0598】 A 96-well microtiter plate (Nunc Maxisorp catalog no. 439454, VWR catalog no. 62409-002) was coated with polyclonal rabbit anti-CHO-PLA2 antibody (1 mg / mL) and then incubated with SuperBlock in TBS (Thermo Scientific catalog no. 37535) to block nonspecific sites. Recombinant PLA2 standard ([PLA-2G15(cg)(34-412)]-6His, 1.05 mg / mL) and drug substance were then added to the plate. The plate was incubated to allow residual PLA2 present in the standard and samples to bind to the polyclonal anti-PLA2 antibody. The plate was washed to remove unbound material, and biotinylated rabbit anti-CHO PLA2 polyclonal antibody (1 mg / mL) was added to the plate. The plate was incubated to allow the biotinylated antibody to bind to the remaining PLA2 antigen bound to the anti-PLA2 antibody. The plate was washed to remove unbound material, and Neutravidin-HRP (enzyme-conjugated horseradish peroxidase, Thermo Scientific Catalog No. 31030) was added to the plate. The plate was incubated to allow the Neutravidin-HRP to bind to the bound biotinylated antibody. The plate was washed to remove unbound material, and K-Blue TMB substrate (Neogen Catalog No. 308177) was added to the plate. The chromogenic substrate was oxidized by the bound enzyme-conjugated antibody, producing a blue color. The reaction was stopped with 4N (2M) sulfuric acid (Ricca Catalog No. 8322-32), resulting in a color change to yellow. The color intensity was directly proportional to the remaining amount of bound PLA2 antigen in the well. The plate was read at 450 nanometers using a plate reader. 【0599】 2. Safety 【0600】 3. Equipment 【0601】 Molecular Devices plate reader or equivalent 【0602】 BioTek ELx405 96W plate washer or equivalent 【0603】 Adjustable pipette with tip, Rainin or equivalent 【0604】 8 or 12 channel pipette with tips, Rainin or equivalent 【0605】 Titer plate shaker, Lab-line or equivalent, room temperature 【0606】 Incubator / shaker, Lab-line Environ plate shaker or equivalent, 37°C 【0607】 pH meter 【0608】 balance device 【0609】 4.Material 【0610】 96-well microtiter plate, Nunc Maxisorp catalog number 439454 (VWR catalog number 62409-002) or equivalent 【0611】 ELISA plate sealing tape Corning catalog number 430454 or equivalent 【0612】 Polypropylene Pipe 【0613】 Milli-Q water, MPS-66 or equivalent 【0614】 Sodium bicarbonate, NaHCO3 FW 84.01g / mol, JT Baker catalog number 3509-01 or equivalent 【0615】 10x Phosphate Buffered Saline with 0.5% Tween-20, Boston BioProducts Catalog No. IBB-171 【0616】 5N Sodium Hydroxide, JT Baker Catalog No. 5671-02 or equivalent 【0617】 5N Hydrochloric Acid, JT Baker Catalog No. 5618-02 or equivalent 【0618】 SuperBlock in TBS, Thermo Scientific catalog number 37535, or equivalent 【0619】 4N sulfuric acid, Ricca catalog number 8322-32 (2N = 1M), or equivalent 【0620】 K-Blue TMB substrate, Neogen catalog number 308177 or equivalent 【0621】 0.22 μM CA sterile filter unit, Corning or equivalent 【0622】 Abcam Sample Diluent, Abcam Catalog Number GR3347356-3; ab193972 【0623】 Rabbit anti-CHO-PLA2 coating polyclonal antibody, 1mg / mL, stored at -80℃ 【0624】 Biotinylated rabbit anti-CHO-PLA2 polyclonal antibody, 1 mg / mL, nominally stored at -80°C 【0625】 Recombinant PLA2 standard, [PLA-2G15(cg)(34-412)]-6His, 1.05 mg / mL, stored at nominal -80°C 【0626】 Neutravidin-HRP conjugate, Thermo Scientific catalog number 31030 or equivalent, aliquoted, stored nominally at 4°C 【0627】 Risankizumab assay control, risankizumab bulk drug substance 145 mg / mL, stored at nominal -80°C 【0628】 5. Preparation of Reagents and Solutions: 【0629】 Note: A reverse pipette was used throughout the assay. Coating buffer and substrate were used cold (removed from a nominal 4°C immediately before use). 【0630】 5.1 50 mM sodium bicarbonate, pH 9.4±0.1 (coating buffer): 【0631】 Add 900 mL of Milli-Q water to a 1 L beaker. 【0632】 Add 4.20 g ± 0.01 g of sodium bicarbonate. 【0633】 Stir until completely dissolved. 【0634】 Adjust the pH to 9.4±0.1 with 5N NaOH and 5N HCl. 【0635】 Transfer to a 1 L volumetric flask and make up to the volume with Milli-Q water. 【0636】 Mix by inversion until homogeneous. 【0637】 Filter through a sterile filter unit (0.22 μm). 【0638】 Store at a nominal temperature of 4°C for a maximum of 7 days from the date of preparation. 【0639】 5.2 Plate Wash Buffer (1x PBS + 0.05% Tween-20): 【0640】 In a 1 L graduated cylinder, add 100 mL of 10× phosphate buffered saline containing 0.5% Tween-20 to 900 mL of Milli-Q water. 【0641】 Stir until homogeneous. 【0642】 Filter through a 0.22 μm sterile filter unit. 【0643】 Store at room temperature for up to 6 months. 【0644】 5.3 Coating antibody mixture: Rabbit anti-CHO PLA2 polyclonal antibody (1 mg / mL), affinity purified: 【0645】 NOTE: Antibody stocks were stored in vials at a nominal -80 °C. Prepare aliquots. One aliquot per plate was removed immediately before use. 50 mM coating buffer (step 5.1) was taken from a nominal 4 °C immediately before use. The mixture was added to the plate while cold. 【0646】 Immediately before use: Dilute coating antibody to a final concentration of 3.0 μg / mL in cold 50 mM sodium bicarbonate as follows: 【0647】 For example, add 36 µL of coating antibody to 11964 µL of cold coating buffer. Mix by gently inverting. 【0648】 5.4 Rabbit anti-CHO PLA2 biotinylated polyclonal antibody (1 mg / mL) 【0649】 Note: Stocks were stored in vials at nominal -80°C. Prepare aliquots: Remove one aliquot per plate at the time of use. 【0650】 Immediately before use: Dilute biotinylated antibody to a final concentration of 0.20 μg / mL in SuperBlock in TBS as follows. 【0651】 For example: Prepare Dilution A by diluting 10 μL of biotinylated antibody with 90 μL of SuperBlock in TBS. Mix by gently inverting. Prepare Dilution B by further diluting 24 μL of Dilution A with 11,976 μL of SuperBlock in TBS (final dilution 1:5000). 【0652】 5.5 Neutravidin-HRP conjugate (1 mg / mL) 【0653】 NOTE: Stocks were stored at a nominal temperature of 4°C. At the time of use, remove one aliquot per plate and allow to warm to room temperature. 【0654】 Immediately before use, mix the conjugate by gently pipetting up and down. Dilute the Neutravidin-HRP conjugate to a final concentration of 0.2 μg / mL. Mix by gently vortexing. 【0655】 For example: Prepare Dilution A by diluting 10 μL of HRP conjugate into 90 μL of SuperBlock in TBS. Mix by gently inverting. Prepare Dilution B by further diluting 24 μL of Dilution A into 11976 μL of SuperBlock in TBS. 【0656】 6. Preparation of Standards and Spikes 【0657】 Note: Stocks were stored in single-use aliquots at nominally -80°C. 【0658】 6.1 Recombinant PLA2 standard, [PLA-2G15(cg)(34-412)]-6His (1.05 mg / mL) 【0659】 6.1.1 Thaw aliquots at room temperature. Serially dilute to a concentration of 21 ng / mL in Abcam Sample Diluent. Serial dilutions for generating the standard curve are shown in the table below using Abcam Sample Diluent in polypropylene tubes. 【0660】 [Table 29] 【0661】 6.1.2 Mix the standards by gently pipetting up and down. 【0662】 6.1.3 Transfer to a polypropylene microtube. 【0663】 6.1.4 Load each standard in triplicate at 100 μL per well into a 96-well microtiter plate. 【0664】 6.2 Spike preparation 【0665】 In a polypropylene microtube, prepare a high-level PLA2 spike of 10.5 ng / mL by diluting the 21,000 ng / mL standard (Standard 1 from step 6.1.1) 2x with Abcam Sample Diluent. 【0666】 For example, dilute 400 μL of 21.000 ng / mL (Standard 1) with 400 μL of Abcam Sample Diluent to a final concentration of 10.5 ng / mL. 【0667】 In a polypropylene microtube, prepare a 5.25 ng / mL mid-level PLA2 spike by diluting the 10.5 ng / mL standard (standard 2 from step 6.1.1) 2x with Abcam sample diluent. 【0668】 In a polypropylene microtube, prepare a 1.313 ng / mL mid-level PLA2 spike by diluting the 2.625 ng / mL standard (standard 4 from step 6.1.1) 2x with Abcam sample diluent. 【0669】 Load triplicate spikes at 100 μL per well into a 96-well microtiter plate. 【0670】 7. Sample Preparation 【0671】 7.1 In a polypropylene tube, dilute risankizumab Process 1 BDS to 2.81 mg / mL in Abcam Sample Diluent (Step 4). Prepare a predilution in a volume sufficient for serial dilutions plated in triplicate at 100 μL / well. 【0672】 NOTE: To prepare spiked samples (step 8) and risankizumab Process 1BDS nominal test dilutions, use the following dilution scheme: 【0673】 [Table 30] 【0674】 7.2 In a polypropylene microtube, further dilute the risankizumab process 1 BDS to 0.18 mg / mL using sample diluent (dilutions 1-4 will be assayed by PLA2 ELISA). 【0675】 [Table 31] 【0676】 7.3 In a polypropylene microtube, dilute Risankizumab Process 2 BDS to 150 mg / mL to 9.38 mg / mL using Sample Diluent. 【0677】 NOTE: To prepare nominal test dilutions for spiked samples (step 8) and Risankizumab Process 2 BDS samples, use the following dilution scheme: 【0678】 [Table 32] 【0679】 7.4 In a polypropylene microtube, further dilute the BDS 150 mg / mL (9.38 mg / mL) solution from Process 2 to 0.59 mg / mL using Abcam sample diluent (dilutions 1-4, assayed by PLA2 ELISA). 【0680】 [Table 33] 【0681】 7.5 Dilute Risankizumab Process 3 BDS 150 mg / mL to 37.5 mg / mL in a polypropylene microfuge tube. 【0682】 NOTE: For spiking of samples in step 8 and for the nominal dilution of BDS samples for Risankizumab Process 3, use the following dilution scheme: 【0683】 [Table 34] 【0684】 7.6 In a polypropylene microtube, further dilute the 37.5 mg / mL Risankizumab Process 3 BDS solution to 2.34 mg / mL using Sample Diluent (Dilutions 1-4, assayed by PLA2 ELISA). 【0685】 [Table 35] 【0686】 7.7 Dilute Risankizumab Process 4 BDS 150 mg / mL to 20 mg / mL in a polypropylene tube. 【0687】 NOTE: To prepare nominal test dilutions for spiked samples (step 8) and risankizumab Process 4 BDS samples, use the following dilution scheme: 【0688】 [Table 36] 【0689】 7.8 In a polypropylene microtube, further dilute the 20 mg / mL risankizumab Process 4 BDS solution to 1.25 mg / mL using sample diluents (dilutions 1-4, assayed by PLA2 ELISA). 【0690】 [Table 37] 【0691】 7.9 Load triplicate wells into the plate at 100 µL / well for each of the nominal test solutions (dilutions 1-4 from steps 7.2, 7.4, 7.6, 7.8). 【0692】 8. Preparation of Spiked Samples 【0693】 8.1 In a polypropylene microtube, mix 400 μL of Abcam Sample Diluent with an equal volume of unspiked risankizumab BDS sample and 400 μL of pre-diluted risankizumab BDS sample (steps 7.1, 7.3, 7.5, and 7.7). 【0694】 8.2 In a polypropylene microfuge tube, mix 400 μL of the 21 ng / mL PLA2 standard solution (Standard 1) with an equal volume of 400 μL of the pre-diluted risankizumab BDS sample (steps 7.1, 7.3, and 7.5 and 7.7). These were spiked with high levels of PLA2 to be tested. 【0695】 8.3 In a polypropylene microfuge tube, mix 400 μL of the 10.5 ng / mL PLA2 standard solution (Standard 2) with an equal volume of 400 μL of the pre-diluted risankizumab BDS sample (steps 7.1, 7.3, and 7.5 and 7.7). These were tested as intermediate-level PLA2 spiked samples. 【0696】 8.4 In a polypropylene microfuge tube, mix 400 μL of the 2.625 ng / mL PLA2 standard solution (Standard 4) with an equal volume of 400 μL of the pre-diluted risankizumab BDS sample (steps 7.1, 7.3, and 7.5 and 7.7). These were tested as low-level PLA2 spiked samples. 【0697】 8.5 The final test concentration of the spiked samples was the same as the unspiked sample preparation (dilutions 1-4) plus 5.250 ng / mL (spiked 2x with the sample diluted from 10.500 ng / mL). 【0698】 8.6 Load triplicate wells onto the plate for unspiked samples and samples spiked with standards (low, medium, and high levels of PLA2) at 100 μL / well for each spiked sample. 【0699】 9. Preparation of Risankizumab Sample Control 【0700】 9.1 A control range must be established for each new control stock solution before use in routine testing. 【0701】 9.2 The control was risankizumab bulk drug substance 145 mg / mL. 【0702】 9.3 Control Stock: Prepare 50 μL aliquots of the batch of risankizumab bulk drug substance and store at a nominal -80°C. 【0703】 Working Control: Thaw an aliquot of control at room temperature. Dilute the control to 7.25 mg / mL in Abcam sample diluent in a polypropylene tube. Transfer the 7.25 mg / mL solution to a polypropylene microtube and load 100 μL per well into 3 wells of the plate. One dilution was adequate. Record the result in the PA control logbook, rounded to the nearest 0.1 ng / mg. 【0704】 [Table 38] 【0705】 10. Preparation of Assay Controls 【0706】 10.1 Prepare a PLA2 standard assay control at 5.25 ng / mL by diluting it using 600 µL of Abcam sample diluent mixed with 200 µL of the 21 ng / mL PLA2 standard solution (standard 1 from step 6.1.1). 【0707】 10.2 A 5.25 ng / mL PLA2 standard solution is used as an assay control and is loaded into triplicate wells at 100 μL / well. 【0708】 11 Steps 【0709】 11.1 Plate Washing Procedure 【0710】 Fill the plate wash bottle with plate wash buffer (1x PBS + 0.05% Tween-20, see step 5.2). Prime the plate washer. Ensure the following parameters: Parameter setting Plate type: 1 Each cycle (total 4 cycles): Volume: 350 μL Immersion time: 10 seconds Suction time: 4 seconds 【0711】 For the BioTek Elx405, the plate washer parameters should be set as follows: Method: 4 cycles Immersion / Shaking: Yes Immersion duration: 0.10 seconds Shake before immersion: No Priming after immersion: No DISP: Dispense volume: 350 μL / well Dispensing flow rate: 05 Dispensing height: 120 (15.240 mm) Horizontal DISP position: 0mm Bottom wash first: No Pre-start Prime: None ASPR: Suction height: 0.29 (3.683 mm) Horizontal ASPR position: -30 (-1.372mm) Suction speed: 03 (4mm / sec) Suction delay: 0004 ms Crosswise ASPR: Yes Crosswise On: All Crosswise height: 0.24 (3.048 mm) Crosswise horizontal position: 30 (1.372 mm) Final suction: Yes Final suction delay: 0000 seconds 【0712】 On each day of use, prime with at least 4 L of Milli-Q water to remove the plate wash buffer. 【0713】 Prime the washer with 1 L of warm 1% Tergazyme, then remove the Tergazyme with at least 4 L of Milli-Q water. Dry and store the washer. 【0714】 11.2 Assay Procedure: A checklist can be used as a guide by checking off the steps completed. In addition, record all equipment used during the assay. 【0715】 NOTE: A reverse pipette was used throughout the assay. Coating buffer and substrate were used cold. Samples, standards, spikes, spiked samples, and controls should be diluted in polypropylene tubes (smallest size possible depending on volume) and transferred to polypropylene microtubes for loading onto the plate. Dilutions may be prepared in polypropylene microtubes, if volume permits. 【0716】 11.2.1 Coat the plate with 100 µL / well of coating antibody (step 5.3). Tap the sides of the plate until the coating solution evenly covers the bottom of the wells, cover with sealing tape, and incubate for 16 hours at a nominal temperature of 4 °C while shaking at 180 rpm on an orbital plate shaker (or equivalent). 【0717】 11.2.2 After incubation, remove the plate and blocking buffer (SuperBlock in PBS) from the refrigerator and allow to equilibrate to ambient temperature. 【0718】 11.2.3 Aspirate the liquid from the plate and tap the plate firmly on a Kimwipe to remove excess buffer. 【0719】 11.2.4 Block the plate by adding 300 μL / well of SuperBlock in TBS to each well. Incubate for 1 hour at 37°C with shaking at 200 rpm on an orbital plate shaker. 【0720】 11.2.5 During the blocking incubation, prepare standards, samples, controls, spikes, and spiked samples (see steps 6-9). Transfer each amount to a polypropylene microtube. Rearrange the tubes to match the plate map. 【0721】 11.2.6 Wash the plate with 350 µL / well of plate wash buffer (step 5.2) using the plate washer settings described in step 11.1 for four wash cycles. Dry the plate on a Kimwipe. 【0722】 11.2.7 Using an 8-channel pipette, add 100 μL / well of standards, samples, controls, spikes, and spiked samples to triplicate wells of the plate. Add 100 μL / well of Abcam sample diluent to all empty wells of the plate to use as blanks. Cover with sealing tape and incubate for 1 hour at 37°C shaking at 200 rpm on an orbital plate shaker (or equivalent). Fill in a template to use as a guide when loading the plate. 【0723】 11.2.8 Wash the plate with 350 µL / well of plate wash buffer (step 5.2) using the plate washer settings in step 11.1 for four wash cycles. Wipe the plate dry on a Kimwipe. 【0724】 11.2.9 Add 100 μL / well of biotinylated antibody (step 5.4) to each well. Cover with sealing tape and incubate the plate for 1 hour at 37°C with shaking at 200 rpm on an orbital plate shaker (or equivalent). 【0725】 11.2.10 Wash the plate with 350 µL / well of plate wash buffer (step 5.2) using the plate washer settings described in step 11.1 for four wash cycles. Dry the plate on a Kimwipe. 【0726】 11.2.11 Add 100 μL / well of Neutravidin-HRP (step 5.5) to each well. Cover with sealing tape and incubate the plate at 37 °C for 1 hour with shaking at 200 rpm on an orbital plate shaker (or equivalent). 【0727】 11.2.12 Wash the plate with 350 µL / well of plate wash buffer (step 5.2) using the plate washer settings in step 11.1 for four wash cycles. Wipe the plate dry on a Kimwipe. 【0728】 11.2.13 Add 100 μL / well of K-Blue substrate to each well. Cover with sealing tape and incubate for 10 minutes at room temperature (25°C ± 2°C) without shaking. 【0729】 11.2.14 Stop the reaction by adding 100 μL / well of 2M (4N) sulfuric acid to each well. 【0730】 11.2.15 Using a plate reader, read the plate at 450 nanometers. 【0731】 11.2.16 Plate reader settings 【0732】 Power on the computer, monitor, and plate reader. 【0733】 Log on to your computer. Double-click the SoftmaxPro icon and select OPEN under the File pull-down menu. 【0734】 Create a new SoftMax file. 【0735】 Verify the parameters and determine the residual phospholipase A2 concentration according to the ELISA protocol parameters. 【0736】 Set up a template and enter the concentrations of the standards. Do not enter the dilution factors for samples, controls, spikes, or spiked samples. Assign the wells containing the diluent as blanks to exclude from all wells (see step 11.2.7). 【0737】 12. Data Analysis and Calculations 【0738】 NOTE: Only samples, spikes, spiked samples, and controls whose optical density fell within the practical limit of quantitation of the standard curve (0.328 ng / mL to 21,000 ng / mL of standard) and met the %CV or %difference criteria described below were accepted. If the sample OD is below the 0.328 ng / mL standard, the result should be reported as less than 0.328 ng / mL. This value should be divided by the diluted risankizumab sample concentration to report the value in ng / mg. If the sample has a high PLA2 concentration, and the unspiked and / or spiked samples are above the standard curve, the value should be reported as greater than 21,000 ng / mL. This value should be divided by the diluted risankizumab sample concentration to report the value in ng / mg. 【0739】 NOTE: If the mean OD value of one sample dilution is equal to or greater than the mean OD value of the 0.328 ng / mL standard and the mean OD value of the other dilution is less than the mean OD value of the 0.328 ng / mL standard, use the value of the dilution with the mean OD value equal to or greater than the mean OD value of the 0.328 ng / mL standard to report the results, provided that the % difference between this ng / mL value and the 0.328 ng / mL value is 30% or less. If the % difference is greater than 30%, repeat the sample. 【0740】 12.1 Standard curve 【0741】 12.1.1 Standard concentrations must be entered into the protocol template. A 4-parameter logistic curve fit was used. 【0742】 12.1.2 The coefficient of determination must be ≥ 0.99 and the %CV between triplicate wells must be ≤ 20%. If these criteria are not met: 【0743】 12.1.2.1 One standard (one level, three wells) may be dropped. If dropping 0.328 ng / mL, only samples and spiked samples with optical densities falling within 0.656 ng / mL and 21.000 ng / mL (remaining standard curve points) are acceptable. 【0744】 12.1.2.2 Additionally, for each standard level triplicate, a single well may be dropped if it is clearly contaminated or shows low binding. When a well is dropped from a standard level, the remaining replicates must have a % difference of 20% or less. 【0745】 12.1.2.3 The %CV of the lowest standard that gives an OD value close to the plate background (blank) must be 30% or less. If one well is dropped, the % difference for the remaining replicates must be 35% or less. If the lowest standard is dropped, only samples and spiked samples whose optical densities fall within the optical densities of the remaining standard curve levels are acceptable. 【0746】 12.1.2.4 Calculate the % Difference as follows: % Difference = (absolute value (OD1-OD2) / mean value) x 100%. 【0747】 If the standards do not meet the above criteria, the assay should be repeated. 【0748】 Report the %CV and / or % difference values of the determination results and the standard curve coefficients. 【0749】 12.2 Sample 【0750】 The %CV between triplicate wells must be ≤ 20%. Report the %CV between triplicate wells. You may drop one well from each sample dilution. The remaining replicates must have a %difference of ≤ 20%. Note: If the OD of the unspiked sample is below the OD of the 0.328 ng / mL standard, the %difference criteria do not apply to the unspiked result. See calculation in step 12.1.2.4. See the first second note in Section 12 for the procedure if one sample dilution is ≥ 0.328 ng / mL (LOQ) and the second dilution is < 0.328 ng / mL. 【0751】 Report the "Unspiked Sample Results" in ng / mL for each dilution. These values were used in the calculation of spike recovery. 【0752】 Calculate the mean of the "Unspiked Sample Results (ng / mL)" and the % difference between dilutions. See calculation in step 12.1.2.4. The % difference between dilutions should be ≤ 20%. Report the results. 【0753】 Calculate PLA2 concentrations (ng / mg) from the mean (ng / mL) values as follows: 【number】 【0754】 Record the results. 【0755】 12.3 Spikes 【0756】 The %CV between triplicate wells should be ≤20%. Record the %CV. You may drop one well from the spike. The %variance for the remaining wells should be ≤20%. See calculation in step 12.1.2.4. 【0757】 PLA2 concentrations are reported in ng / mL and were used to calculate spike recovery. 【0758】 The resulting spike concentration (ng / mL) should be ±20% of the theoretical spike concentration. The results are recorded and indicated as pass or fail. If the spike result is not within 20% of the theoretical value, the assay should be repeated. 【number】 【0759】 Spiked samples 【0760】 The %CV between triplicate wells must be ≤20%. Record the %CV. You may drop one well from each spiked sample dilution. The remaining replicates must have a % difference of ≤20%. See calculation in step 12.1.2.4. 【0761】 Report the "Spiked Sample Results" in ng / mL for each dilution. Record the % difference between the duplicate dilutions (see step 12.1.2.4 for the formula). The % difference between the dilutions should be 25% or less. These results were used in the calculation of spike recovery. 【0762】 Calculate the % spike recovery for each dilution set using the following formula: 【number】 【0763】 Notes: (1) If the OD of the unspiked sample is below the standard (LOQ) of 0.328 ng / mL, consider the value as zero in calculating % spike recovery. 【0764】 The % spike recovery should be 100% ± 50% (50% to 150%) for each dilution of each sample. Record the results and pass / fail. 【0765】 12.5 Control 【0766】 12.5.1 The %CV between triplicate wells must be ≤20%. Record the %CV results. You may drop one well from the control. The remaining replicates must have a % difference of ≤20%. See calculation in step 12.1.2.4. 【0767】 12.5.2 Record the PLA2 concentration in ng / mL. 【0768】 Calculate the PLA2 concentration (ng / mg) as follows: 【number】 【0769】 Report results appropriately and in the control logbook (ng / mg). If the controls are outside the set range, the assay should be repeated. 【0770】 12.6 Blanks 【0771】 If any blank wells show significant contamination, mask the well(s). 【0772】 12.7 Assay Range 【0773】 The assay range should be determined to be 0.328 ng / mL to 21.000 ng / mL. 【0774】 13. Protocols for Preparation of Recombinant PLA2, Polyclonal Rabbit Anti-CHO PLA2 Antibody, and Purification of PLA2 Antibody 【0775】 13.1 Preparation of PLA2G15 antigen 【0776】 The DNA sequence encoding PLA2G15 from Cricetulus griceus (Chinese hamster) (UNIPROT G3HKV9; amino acids 1–412) was synthesized and cloned into the pHybE (U.S. Patent No. 8,187,836 B2) vector, incorporating an in-frame hexahistidine tag (SEQ ID NO: 15). The pHybE expression vector utilizes the EF-1α promoter and the OriP origin of replication from the Epstein-Barr virus (EBV). 【0777】 This plasmid was transfected into CHO-3E7 cells (NRC Canada) grown at 3.3 x 10e6 cells / ml in BalanCD CHO medium (Irvine Scientific) at a PEI:DNA ratio of 8:1 using the transfection reagent Polyethylenimine Max (PEI Max, Polysciences Inc.). 24 hours after transfection, the transfected cell culture was fed with 4% 1x CHO4 feed (Irvine Scientific), 5% Transfectly Supplement (Irvine Scientific), and 2.5 g / L glucose. Seven days after transfection, the transfected cell culture was clarified by centrifugation followed by filtration through a Sartopore-2 0.45 + 0.2 mm filter (Sartorius). 【0778】 The clarified medium was loaded onto a 5 ml HisTrap Excel column (Cytiva) equilibrated with PBS, pH 7.4. The column was washed with 25 mM imidazole in PBS, pH 7.4, and bound protein was eluted with 500 mM imidazole in PBS, pH 7.4. The eluted protein was concentrated using a Centricon plus-70 centrifugal filter device (Millipore) with a 30 kDa molecular weight cutoff, and further purified by SEC on a 26 / 60 Superdex 200 column (Cytiva) equilibrated and run in PBS, pH 7.4. Fractions containing PLA2G15 were pooled, the concentration was determined by absorbance at 280 nm, and the sample was analyzed by SEC, SDS-PAGE, and mass spectrometry. Purified [PLA2G15(cg)(34-412)]-6His was stored in aliquots at -80°C. 【0779】 13.2. Production of Polyclonal Rabbit Anti-CHO PLA2 Antibodies 【0780】 Polyclonal rabbit anti-CHO PLA2 antibodies can be produced by immunizing rabbits (eg, New Zealand White rabbits) with the PLA2G15 antigen described above. 【0781】 The antigen may be used with an adjuvant (eg, Freund's adjuvant) to enhance the immune response for polyclonal antibody production. 【0782】 The antigen may be injected into the animal intramuscularly, intradermally, or subcutaneously. 【0783】 Booster immunizations can be administered, for example, 1 to 8 weeks after the priming immunization and can be continued at intervals of 1 to 4 weeks. 【0784】 Polyclonal antibody production in rabbits can be assessed by taking serum samples before the priming immunization and after each of the priming and booster immunizations. 【0785】 Polyclonal antibody production may be terminated when antibody titers reach acceptable levels. 【0786】 The animals are exsanguinated and serum is collected from the whole blood. 【0787】 13.3. Affinity Purification of Polyclonal Rabbit Anti-CHO PLA2 Antibody 【0788】 PBS-equilibrated PLA2-conjugated CnBr-Sepharose beads (number 17-0430-01) were added to 740 ml of anti-PLA2 serum and incubated at 4°C with rotation for 2 days. 【0789】 An EconoPac column or similar was used to expel the beads and the flow-through was collected and retained for control purposes. 【0790】 The beads were washed with approximately 200 ml of TTBS and approximately 75 ml of Gentle Elution Buffer and 200 ml of TTBS, and eluted with approximately 50 ml of IgG Elution Buffer (Pierce #21009). 【0791】 The eluate was neutralized with 10% v / v 2M Tris-HCl pH 7.5 (pre-dispensed in collection tubes). Protein peaks were measured using a 1:5 water diluted Bradford solution pre-dispensed in a 96-well plate (200 ul Bradford / well, add 20 ul of eluate). 【0792】 PLA2-bound CNBr-Sepharose beads were neutralized with 10 bed volumes of PBS and added back to the flow-through at 4°C over the weekend between the bead draining step (using EconoPac) and the second round of incubation. 【0793】 The eluate was dialyzed three times against PBS exchange buffer on a Big Tuna (Unchained Labs) (50ml of eluate was subjected to a 96% exchange into PBS on a Big Tuna. After the first round, 25ml was removed, mixed, the concentration measured and the pool divided into 96x aliquots at 250ul / aliquot. The remaining 25ml was transferred to a new Big Tuna filter plate and subjected to a new round of 96% exchange into a final 2x concentration of PBS, followed by biotinylation). 【0794】 PLBL2 ELISA 【0795】 1. Principles 【0796】 A 96-well microtiter plate (Nunc Maxisorp catalog no. 439454, VWR catalog no. 62409-002) was coated with polyclonal anti-PLBL2 antibody (2 mg / mL). The plate was then incubated with SuperBlock (Thermo Scientific catalog no. 37515) in PBS to block nonspecific sites. Recombinant PLBL2 standard ([PLBL-2(cg)(38-585)]-6His, 1.27 mg / mL) and drug substance were then added to the plate. The plate was incubated to allow the remaining PLBL2 present in the standard and samples to bind to the polyclonal anti-PLBL2 antibody. The plate was washed to remove unbound material, and biotinylated anti-PLBL2 polyclonal antibody (1 mg / mL) was added to the plate. The plate was then incubated to allow the biotinylated antibody to bind to the remaining PLBL2 antigen bound to the anti-PLBL2 antibody. The plate was washed to remove unbound material, and streptavidin-poly-HRP (enzyme-conjugated horseradish peroxidase; Thermo Scientific catalog no. 21140) was added to the plate. The plate was incubated to allow the streptavidin-poly-HRP to bind to the conjugated biotinylated antibody. The plate was washed to remove unbound material, and K Blue TMB substrate (Neogen catalog no. 308177) was added to the plate. The chromogenic substrate was oxidized by the bound enzyme-conjugated antibody, producing a blue color. The reaction was stopped with 4N (2M) sulfuric acid (Ricca catalog no. 8310-32), resulting in a color change to yellow. The color intensity was directly proportional to the amount of remaining PLBL2 antigen bound to the well. The plate was read at 450 nanometers using a plate reader. 【0797】 2. Safety 【0798】 Standard laboratory safety precautions. 【0799】 3. Equipment 【0800】 Molecular Devices plate reader or equivalent 【0801】 Tecan 96W plate washer or equivalent 【0802】 Adjustable pipette with tip, Rainin or equivalent 【0803】 8 or 12 channel pipette with tips, Rainin or equivalent 【0804】 Titer plate shaker, Lab-line or equivalent, room temperature 【0805】 Incubator / shaker, Lab-line Environ plate shaker or equivalent, room temperature 【0806】 pH meter 【0807】 balance device 【0808】 4.Material 【0809】 96-well microtiter plate, Nunc Maxisorp catalog number 439454 (VWR catalog number 62409-002) or equivalent 【0810】 ELISA plate sealing tape - Corning catalog number 430454 or equivalent 【0811】 Polypropylene Tube 【0812】 Opaque Microtiter Plate Cover 【0813】 Milli-Q water, MPS-66 or equivalent 【0814】 Sodium bicarbonate, NaHCO3 FW 84.01g / mol, JT Baker catalog number 3509-01 or equivalent 【0815】 Tween-20, JT Baker catalog number X251-07 or equivalent 【0816】 5N Sodium Hydroxide, JT Baker Catalog No. 5671-02 or equivalent 【0817】 5N Hydrochloric Acid, JT Baker Catalog No. 5618-02 or equivalent 【0818】 Sodium chloride, NaCl SW 58.44 g / mol, Sigma catalog number S3014 or equivalent 【0819】 Heptahydrate dibasic sodium phosphate, crystalline Na2HPO4*7H2O, FW 268.07, JT Baker catalog number 3817-01 or equivalent 【0820】 SuperBlock in PBS, Thermo Scientific catalog number 37515 or equivalent 【0821】 4N sulfuric acid, Ricca catalog number 8310-32 (2N = 1M) or equivalent 【0822】 K-Blue TMB substrate, Neogen catalog number 308177 or equivalent 【0823】 0.22 μM CA sterile filter unit, Corning or equivalent 【0824】 Plate wash buffer, 1x PBS + 0.05% Tween-20, MPS-40, store at room temperature 【0825】 Anti-id-PLBL2 polyclonal coating antibody, 2mg / mL, stored at nominal -80°C 【0826】 Biotinylated anti-PLBL2 polyclonal detection antibody, 1 mg / mL, nominally stored at -80°C 【0827】 Recombinant PLBL2 standard, [PLBL-2(cg)(38-585)]-6His, 1.27 mg / mL, stored at nominal -80°C 【0828】 Streptavidin-poly-HRP conjugate, Thermo Scientific catalog number 21140 or equivalent, aliquoted, stored at nominal 4°C 【0829】 PLBL2 Assay Control, Risankizumab Ultrafiltration / Dialysis (UF / DF) Retentate 10 mg / mL, Lot No. 91400096, Aliquots, Store Nominally at -80°C 【0830】 5. Preparation of Reagents and Solutions: 【0831】 Note: A reverse pipette was used throughout the assay. Coating buffer and substrate were used cold (removed from a nominal 4°C immediately before use). 【0832】 5.1 50 mM sodium bicarbonate, pH 9.4±0.1 (coating buffer): 【0833】 Add 900 mL of Milli-Q water to a 1 L beaker. 【0834】 Add 4.20 g ± 0.01 g of sodium bicarbonate. 【0835】 Stir until completely dissolved. 【0836】 Adjust the pH to 9.4±0.1 with 5N NaOH and 5N HCl. 【0837】 Transfer to a 1 L volumetric flask and make up to the volume with Milli-Q water. 【0838】 Mix by inversion until homogeneous. 【0839】 Filter through a sterile filter unit (0.22 μm). 【0840】 Store at a nominal temperature of 4°C for a maximum of 7 days from the date of preparation. 【0841】 5.2 10x Phosphate Buffered Saline (PBS), MPS-73: 【0842】 Add 800 mL of Milli-Q water to a glass beaker. 【0843】 Add 80.0 g of NaCl to a final concentration of 80.0 g / L or 1.37M. 【0844】 Add 2.00 g of KCl to a final concentration of 2.00 g / L or 0.0268M. 【0845】 27.88g Na2HPO4·7H2O / Add 100 mL of PBS to a final concentration of 0.1040 M, or 0.1040 M. 【0846】 Add 2.40 g of KH2PO4 to a final concentration of 2.40 g / L, or 0.0176M. 【0847】 Add Milli-Q water to make 1000 mL. 【0848】 Stir until homogeneous. 【0849】 Check the pH and, if necessary, adjust it to 6.8-6.9 using 5N HCl. 【0850】 Stir until homogeneous. 【0851】 Sterilize at 123°C for 30 minutes. 【0852】 Store at room temperature for up to 12 months. 【0853】 5.3 Plate Wash Buffer / Assay Diluent / MPS-40 (1x PBS + 0.05% Tween-20): 【0854】 In a 1 L graduated cylinder, add 100 mL of MPS-73 (step 5.2) to 900 mL of Milli-Q water. 【0855】 0.5 mL of tween-20 is added to the solution. 【0856】 Stir until homogeneous. 【0857】 Check the pH and adjust if necessary to 7.40±0.05 with 5N HCl. 【0858】 Stir until homogeneous. 【0859】 Filter through a 0.22 μm sterile filter unit. 【0860】 Store at room temperature for up to 6 months. 【0861】 5.4 Coating antibody mixture: anti-idPLBL2 polyclonal antibody (2 mg / mL), affinity purified: 【0862】 NOTE: Antibody stocks were stored in vials at a nominal -80 °C. Prepare aliquots. Remove one aliquot per plate immediately before use. Take 50 mM coating buffer (step 5.1) from a nominal 4 °C immediately before use. Add the mixture to the plate while cold. 【0863】 Immediately before use: Dilute coating antibody to a final concentration of 1 μg / mL in cold 50 mM sodium bicarbonate as follows: 【0864】 For example: Add 6 μL of coating antibody to 11994 μL of cold coating buffer. Mix by gently inverting. 【0865】 5.5 Anti-PLBL2 polyclonal antibody-biotin conjugate (1 mg / mL) 【0866】 NOTE: Stocks were stored in vials at nominal -80°C. Prepare aliquots. Take one aliquot per plate at the time of use. 【0867】 Immediately before use: Dilute biotinylated antibody to a final concentration of 0.80 µg / mL in MPS-40 as follows (step 5.3): 【0868】 For example, dilute 10 μL of biotinylated antibody in 12490 μL of MPS-40. Mix by gently inverting. 【0869】 5.6 Streptavidin-poly-HRP conjugate (0.5 mg / mL) 【0870】 NOTE: Stocks were stored at a nominal temperature of 4°C. At the time of use, remove one aliquot per plate and warm to room temperature. 【0871】 Immediately before use, mix the conjugate by gently pipetting up and down. Dilute the streptavidin-poly-HRP conjugate to a final concentration of 0.083 μg / mL. Mix by gently vortexing. 【0872】 For example: Dilute 10 μL of HRP conjugate with 990 μL of MPS-40 (Diluent A). Then further dilute the HRP conjugate by adding 200 μL of Diluent A to 11,800 μL of MPS-40 (Diluent B). 【0873】 6. Preparation of Standards and Spikes 【0874】 Note: Stocks were stored in single-use aliquots at nominally -80°C. 【0875】 6.1 Recombinant PLBL2 standard, [PLBL-2(cg)(38-585)]-6His (1.27 mg / mL) 【0876】 6.1.1 Thaw an aliquot at room temperature. Serial dilutions are performed in MPS-40 (step 5.3) down to a concentration of 4 ng / mL. The serial dilutions for generating the standard curve are shown in the table below using MPS-40 in polypropylene tubes. 【0877】 [Table 39] 【0878】 6.1.2 Mix the standards by gently pipetting up and down. 【0879】 6.1.3 Transfer to a polypropylene microtube. 【0880】 6.1.4 Load each standard in triplicate at 100 μL per well into a 96-well microtiter plate. 【0881】 6.2 Spike preparation 【0882】 6.2.1 In a polypropylene microtube, prepare a 0.320 ng / mL recombinant PLBL2 spike from the 0.640 ng / mL standard (standard 3) prepared above by diluting 2x in MPS-40 (step 5.3). Make one dilution. 【0883】 6.2.2 For example, dilute 300 μL of 0.640 ng / mL (Standard 3) with 300 μL of MPS-40 to a final concentration of 0.320 ng / mL. 【0884】 6.2.3 Load spikes in triplicate at 100 μL per well into a 96-well microtiter plate. 【0885】 6.2.4 Use the 0.640 ng / mL standard (standard 3) from step 6.1.1 to spike the samples. 【0886】 7. Sample Preparation 【0887】 7.1 In a polypropylene tube, dilute BDS to 0.045 mg / mL in MPS-40 (step 5.3). Make a predilution in a volume sufficient for serial dilutions plated in triplicate at 100 µL per well. 【0888】 NOTE: To prepare spiked samples (step 8) and BDS nominal test dilutions, use the following dilution scheme: 【0889】 [Table 40] 【0890】 7.2 In a polypropylene microtube, further dilute the 0.1788 mg / mL solution to 0.01117 mg / mL in MPS-40 (step 5.3). 【0891】 [Table 41] 【0892】 7.3 Risankizumab clarification harvest followed the same dilution scheme as risankizumab BDS, with final plate serial dilutions of 1000x, 2000x, 4000x, and 8000x. 【0893】 NOTE: Use the following dilution scheme to prepare the spiked samples (step 8) and clarified harvested nominal test dilutions. 【0894】 [Table 42] 【0895】 7.4 In a polypropylene microtube, further dilute the 0.00098 mg / mL (0.98 μg / mL) solution to 0.061 μg / mL in MPS-40 (step 5.3). 【0896】 [Table 43] 【0897】 7.5 Risankizumab Protein A eluate followed the same dilution scheme as Risankizumab BDS, with final plate serial dilutions of 1000x, 2000x, 4000x, and 8000x. 【0898】 NOTE: For spiking samples in step 8 and preparing nominal test dilutions of Protein A eluate, use the following dilution scheme: 【0899】 [Table 44] 【0900】 7.6 In a polypropylene microtube, further dilute the 0.0165 mg / mL solution to 0.00103 mg / mL in MPS-40 (step 5.3). 【0901】 [Table 45] 【0902】 7.7 For each of the nominal test solutions, load triplicate wells at 100 μL per well on the plate for a total of 36 wells. 【0903】 8. Preparation of Spiked Samples 【0904】 8.1 In a polypropylene microtube, add 400 μL of the 0.640 ng / mL standard (Standard 3) to an equal number of new microtubes as unspiked sample test diluents. 【0905】 8.2 Transfer 400 µL of the 500x predilutions (i.e., 0.1788 mg / mL BDS, 0.00098 mg / mL clarified harvest, and 0.0165 mg / mL Protein A eluate, respectively) from steps 7.1, 7.3, and 7.5 to a new microtube containing 400 µL of 0.640 ng / mL spike. This was tested as spike dilution 1. 【0906】 8.3 Dilute dilutions 1, 2, and 3 from each of the unspiked sample preparations 2x by adding 400 µL of sample to a new microtube containing 400 µL of 0.640 ng / mL spike (steps 7.2, 7.4, and 7.6). These were tested as spiked dilutions 2-4. 【0907】 For example, 89.38 mg / mL risankizumab BDS is spiked to final test dilutions of 1000×, 2000×, 4000×, 8000×. 【0908】 [Table 46] 【0909】 8.4 Final test concentrations for spiked samples were similar to the unspiked sample preparations (dilutions 1-4) + 0.320 ng / mL (spike diluted 2x in sample from 0.640 ng / mL). 【0910】 8.5 Load triplicate wells of each spiked sample solution onto the plate at 100 μL per well for a total of 36 wells. 【0911】 9. Preparation of Controls 【0912】 9.1 A control range must be established for each new control stock solution before use in routine testing. 【0913】 The control was risankizumab UF / DF retentate 10 mg / mL. 【0914】 9.2 Control Stock: Prepare 500 μL aliquots of the batch of risankizumab UF / DF retentate and store at nominally -80°C. 【0915】 9.3 Working Controls: Thaw an aliquot of control at room temperature. In a polypropylene tube, dilute the control to 1 mg / mL with MPS-40 (step 5.3). Transfer the 1 mg / mL solution to a polypropylene microtube and load 100 μL per well into a 3-well plate. One dilution was adequate. Record the result in the PA control logbook, rounded to the nearest 0.1 pg / mg. 【0916】 [Table 47] 【0917】 10. Procedure 【0918】 10.1 Plate Washing Procedure 【0919】 Fill the plate wash bottle with plate wash buffer (see step 5.3, MPS-40). Prime the plate washer. Check the following parameters: Parameter setting Plate type: 1 Each cycle (total 4 cycles): Volume: 300 μL Immersion time: 10 seconds Suction time: 4 seconds 【0920】 For the BioTek Elx405, the plate washer parameters should be set as follows: Method: 4 cycles Immersion / Shaking: Yes Immersion duration: 0.10 seconds Shake before immersion: No Priming after immersion: No DISP: Dispense volume: 300 μL / well Dispensing flow rate: 05 Dispensing height: 120 (15.240 mm) Horizontal DISP position: 0mm Bottom wash first: No Pre-start Prime: None ASPR: Suction height: 0.29 (3.683 mm) Horizontal ASPR position: -30 (-1.372mm) Suction speed: 03 (4mm / sec) Suction delay: 0004 ms Crosswise ASPR: Yes Crosswise On: All Crosswise height: 0.24 (3.048 mm) Crosswise horizontal position: 30 (1.372 mm) Final suction power: Yes Final suction delay: 0000 seconds 【0921】 On each day of use, prime with at least 4 L of Milli-Q water to remove the plate wash buffer. 【0922】 Prime the washer with 1 L of warm 1% Tergazyme, then remove the Tergazyme with at least 4 L of Milli-Q water. Dry and store the washer. 【0923】 10.2 Assay Procedure: A checklist can be used as a guide by checking off the steps completed. In addition, record all equipment used during the assay. 【0924】 NOTE: A reverse pipette was used throughout the assay. Coating buffer and substrate were used cold. Samples, standards, spikes, spiked samples, and controls should be diluted in polypropylene tubes (smallest size possible depending on volume) and transferred to polypropylene microtubes for loading onto the plate. Dilutions may be prepared in polypropylene microtubes, if volume permits. 【0925】 10.2.1 Coat the plate with 100 μL / well of coating antibody (step 5.4). Tap the sides of the plate until the coating solution evenly covers the bottom of the wells, cover with sealing tape, and incubate for 18 ± 1 h at a nominal 4°C while shaking at 180 rpm on an orbital plate shaker (or equivalent). 【0926】 10.2.2 After incubation, remove the plate and blocking buffer (SuperBlock in PBS) from the refrigerator and allow to equilibrate to ambient temperature. 【0927】 10.2.3 Drain the liquid from the plate into a sink and tap the plate firmly on a Kimwipe to remove excess buffer. 【0928】 10.2.4 Block the plate by adding 300 μL / well of SuperBlock in PBS to each well. Incubate at room temperature without shaking for 1 hour. 【0929】 10.2.5 During the blocking incubation, prepare standards, samples, controls, spikes, and spiked samples (see steps 6-9). Transfer each amount to a polypropylene microtube. Rearrange the tubes to match the plate map. 【0930】 10.2.6 Wash the plate using 300 μL / well with the MPS-40 washer (step 5.3) for four wash cycles. Dry the plate on a Kimwipe. 【0931】 10.2.7 Using an 8-channel pipette, add 100 μL / well of standards, samples, controls, spikes, and spiked samples to triplicate wells of the plate. Add 100 μL / well of MPS-40 to all empty wells of the plate (step 5.3) to serve as blanks. Cover with sealing tape and incubate for 2 hours at room temperature (25°C ± 2°C) with shaking at 400 rpm on an orbital plate shaker (or equivalent). Fill in a template to use as a guide when loading the plate. 【0932】 10.2.8 Wash the plate using 300 μL / well for four wash cycles using the plate washer MPS-40 (step 5.3). Blot the plate on a Kimwipe. 【0933】 10.2.9 Add 100 μL / well of biotinylated antibody (step 5.5) to each well. Protect the reaction from light by covering with sealing tape and an opaque plate cover. Incubate the plate at room temperature (25°C ± 2°C) for 45 minutes, shaking at 400 rpm on an orbital plate shaker (or equivalent). 【0934】 10.2.10 Wash the plate using 300 μL / well for four wash cycles using the plate washer MPS-40 (step 5.3). Dry the plate on a Kimwipe. 【0935】 10.2.11 Add 100 µL / well of streptavidin-poly-HRP (step 5.6) to each well. Cover with sealing tape and an opaque plate cover to protect the reaction from light. Incubate the plate for 30 minutes at room temperature (25°C ± 2°C) while shaking at 400 rpm on an orbital plate shaker (or equivalent). 【0936】 10.2.12 Wash the plate using 300 μL / well for four wash cycles using the plate washer MPS-40 (step 5.3). Dry the plate on a Kimwipe. 【0937】 10.2.13 Add 100 μL / well of K-Blue substrate to each well. Protect the reaction from light by covering with sealing tape and an opaque plate cover. Incubate for 10 minutes at room temperature (25°C ± 2°C) with shaking at 400 rpm on an orbital plate shaker (or equivalent). 【0938】 10.2.14 Stop the reaction by adding 100 μL / well of 2M (4N) sulfuric acid to each well. 【0939】 10.2.15 Using a plate reader, read the plate at 450 nanometers. 【0940】 10.2.16 Plate reader settings 【0941】 Power on the computer, monitor, and plate reader. 【0942】 Log on to your computer. Double-click the Softmax Pro icon and select OPEN under the File pull-down menu. 【0943】 Create a new SoftMax file and set the parameters and plate map. 【0944】 Verify the parameters and determine the residual phospholipase B-like 2 concentration according to the ELISA protocol parameters. 【0945】 Set up the template and enter the concentrations of the standards. Do not enter the dilution factors for samples, controls, spikes, or spiked samples. Assign the wells containing the diluent as blanks to exclude from all wells (see step 10.2.7). 【0946】 11. Data Analysis and Calculations 【0947】 NOTE: Only samples, spikes, spiked samples, and controls whose optical density fell within the practical limit of quantitation of the standard curve (0.041 ng / mL to 4 ng / mL of standard) and met the %CV or %difference criteria described below were accepted. If the sample OD is below the 0.041 ng / mL standard, the result should be reported as less than 0.041 ng / mL. This value should be divided by the diluted sample concentration and multiplied by 1000 to report the value in pg / mg. If the sample has a high PLBL2 concentration, and the unspiked and / or spiked samples are above the standard curve, the value should be reported as greater than 4 ng / mL. This value should be divided by the diluted sample concentration and multiplied by 1000 to report the value in pg / mg. If a sample is below the 0.041 ng / mL standard, the sample value should be considered zero for the spike recovery calculation. 【0948】 NOTE: If one sample dilution has a mean OD equal to or greater than that of the 0.041 ng / mL standard and the other has a mean OD less than that of the 0.041 ng / mL standard, report the result using the value of the dilution with a mean OD greater than or equal to that of the 0.041 ng / mL standard, provided that the % difference between this ng / mL value and the 0.041 ng / mL value is 30% or less. If the % difference is greater than 30%, repeat the sample. 【0949】 11.1 Standard curve 【0950】 11.1.1 Standard concentrations must be entered into the protocol template. A 4-parameter logistic curve fit was used. 【0951】 11.1.2 The coefficient of determination must be ≥ 0.99 and the %CV between triplicate wells must be ≤ 20%. If these criteria are not met: 【0952】 11.1.2.1 One standard (one level, three wells) may be dropped. If dropping 0.016 ng / mL, only samples and spiked samples with optical densities falling within 0.041 ng / mL and 4 ng / mL (remaining standard curve points) are acceptable. 【0953】 11.1.2.2 Additionally, for each standard level triplicate, a single well may be dropped if it is clearly contaminated or shows low binding. When a well is dropped from a standard level, the remaining replicates must have a % difference of 20% or less. 【0954】 11.1.2.3 The %CV for the lowest standard, which shows an OD value close to the plate background (blank), must be 30% or less. If one well is dropped, the % difference for the remaining replicates must be 35% or less. If the lowest standard is dropped, only samples and spiked samples whose optical densities fall within the optical densities of the remaining standard curve levels are acceptable. 【0955】 11.1.2.4 Calculate the % Difference as follows: % Difference = (absolute value (OD1-OD2) / mean value) x 100%. 【0956】 11.1.3 If the reference material does not meet the above criteria, the assay should be repeated. 【0957】 11.1.4 Report the %CV and / or % difference values of the determination results and the standard curve coefficients. 【0958】 11.2 Sample 【0959】 The %CV between triplicate wells must be ≤ 20%. Report the %CV between triplicate wells. You may drop one well from each sample dilution. The remaining replicates must have a %difference of ≤ 20%. Note: If the OD of the unspiked sample is below the OD of the 0.041 ng / mL standard, the %difference criteria do not apply to the unspiked result. See calculation in step 11.1.2.4. See the first second note in Section 11 for the procedure if the first sample dilution is ≥ 0.041 ng / mL (LOQ) and the second dilution is < 0.041 ng / mL. 【0960】 Report the "Unspiked Sample Results" in ng / mL for each dilution. These values were used in the calculation of spike recovery. 【0961】 Calculate the mean of the "Unspiked Sample Results (ng / mL)" and the % difference between dilutions. See calculation in step 11.1.2.4. The % difference between dilutions should be 25% or less. Report the results. 【0962】 Calculate the PLBL2 concentration (ng / mL) from the mean (ng / mL) values as follows: 【number】 【0963】 Record the results. 【0964】 11.3 Spikes 【0965】 The %CV between triplicate wells should be ≤20%. Record the %CV. You may drop one well from the spike. The %variance for the remaining wells should be ≤20%. See calculation in step 11.1.2.4. 【0966】 PLBL2 concentrations are reported in ng / mL and were used to calculate spike recovery. 【0967】 The resulting spike concentration (ng / mL) should be ±20% of the theoretical spike concentration. The results are recorded and indicated as pass or fail. If the spike result is not within 20% of the theoretical value, the assay should be repeated. 【number】 【0968】 11.4 Spiked Samples 【0969】 The %CV between triplicate wells must be ≤20%. Record the %CV. You may drop one well from each spiked sample dilution. The remaining replicates must have a % difference of ≤20%. See calculation in step 11.1.2.4. 【0970】 Report the "Spiked Sample Results" in ng / mL for each dilution. Record the % difference between the duplicate dilutions (see step 11.1.2.4 for the formula). The % difference between the dilutions should be 25% or less. These results were used in the calculation of spike recovery. 【0971】 Calculate the % spike recovery for each dilution set using the following formula: 【number】 【0972】 Notes: (1) If the OD of the unspiked sample is below the standard (LOQ) of 0.041 ng / mL, consider the value as zero in calculating the % spike recovery. 【0973】 The % recovery of the spike should be 100% ± 50% (50% to 150%) for each dilution of each sample. Record the results and pass / fail. 【0974】 11.5 Control 【0975】 The %CV between triplicate wells must be 20% or less. Record the %CV results. You may drop one well from the control. The remaining replicates must have a % difference of 20% or less. See calculation in step 11.1.2.4. 【0976】 Record the PLBL2 concentration in ng / mL. 【0977】 Calculate the PLBL2 concentration (ng / mg) as follows: 【number】 【0978】 Report results appropriately and in the control logbook (pg / mg). If the controls are outside the established range, the assay should be repeated. 【0979】 11.6 Blanks 【0980】 If any blank wells show significant contamination, mask the well(s). 【0981】 11.7 Assay Range 【0982】 The assay range should be determined to be 0.041 ng / mL to 4.000 ng / mL. 【0983】 LPL ELISA 【0984】 1. Principles 【0985】 A commercially available kit was used to perform this assay: Mouse LPL / Lipoprotein Lipase ELISA Kit (catalog number LS-F11957; Lifespan Bioscience (LSBio)). The 96-well plate of the purchased kit was precoated with anti-LPL antibody and blocked with blocking reagent. Mouse LPL standards (provided with the kit) and risankizumab samples (serial diluted using the sample diluent provided with the kit) were added to the 96-well plate. The 96-well plate was incubated to allow LPL present in the standards and samples to bind to the plate-bound polyclonal anti-LPL antibody. The 96-well plate was washed to remove unbound material, and biotinylated anti-LPL antibody (detection reagent A) was added to the plate. The plate was incubated to allow the biotinylated antibody to bind to the LPL antigen that binds to the anti-LPL antibody. The plate was washed to remove unbound material, and streptavidin-HRP conjugate (enzyme-labeled horseradish peroxidase, detection reagent B) was added to the plate. The plate was incubated to allow the streptavidin-HRP to bind to the bound biotinylated antibody. The plate was washed to remove unbound material, and TMB substrate was added to the plate. The chromogenic substrate TMB was oxidized by the bound enzyme (HRP)-conjugated antibody, producing a blue color. The colorimetric reaction was stopped with the provided stop solution, resulting in a color change to yellow. The optical density of each well was directly proportional to the amount of CHO LPL antigen bound to the well. The plate was read at 450 nm within 2 minutes after adding the stop solution. Blank subtraction was performed. 【0986】 2. Safety 【0987】 Standard laboratory safety precautions. 【0988】 3. Equipment 【0989】 Molecular Devices plate reader or equivalent 【0990】 Reporter pipette, Eppendorf or equivalent 【0991】 Sterile filter unit (0.2 μm) 【0992】 Adjustable pipette with tip, Rainin or equivalent 【0993】 8 or 12 channel pipette with tips, Rainin or equivalent 【0994】 Titer plate shaker, Wallac Delfia Cat. No. 1296-004 (1.5 mm shaking orbit) or equivalent, room temperature, speed approximately 350 RPM 【0995】 pH meter 【0996】 balance device 【0997】 4.Material 【0998】 ELISA sealing tape - Corning catalog number 430454 or equivalent 【0999】 Polypropylene Microtubes 【1000】 Immunoware microtubes with rack, 1.1 ml or equivalent 【1001】 Low protein binding tubes, Eppendorf catalog numbers 022431064 (0.5 mL), 022431081 (1.5 mL), and 022431102 (2.0 mL), and 0030108302 (5.0 mL) 【1002】 MilliQ Water 【1003】 0.22 μM CA sterile filter unit, Corning or equivalent 【1004】 Mouse LPL / Lipoprotein Lipase ELISA Kit, Lifespan Bioscience Catalog No. LS-F11957, each kit contains: 【1005】 96-well ELISA plates (pre-treated with anti-LPL antibody coating and blocked with blocking buffer provided by the supplier) 【1006】 Mouse LPL standard (lyophilized), 2 vials (depending on the lot number; lots from 2019 have two vials of 4 ng / vial lyophilized stock, and lots from 2021 have two vials of 1 ng / vial lyophilized stock) 【1007】 Sample diluent (20 mL) 【1008】 Biotinylated anti-LPL antibody (capture antibody) - Detection Reagent A (120 μL) 【1009】 Assay Diluent A (less than 10 mL): 【1010】 Streptavidin-HRP conjugate - Detection Reagent B (120 μL) 【1011】 Assay Diluent B (less than 10 mL): 【1012】 Wash buffer (25x concentration), 30 mL 【1013】 TMB substrate, 10ML 【1014】 Stop solution, 10mL 【1015】 Adhesive Plate Sealer 【1016】 5. Preparation of Reagents and Solutions: 【1017】 Note: Reverse pipetting was used throughout the assay unless otherwise stated. All buffers were used at room temperature. 【1018】 5.1 1x Plate Wash Buffer 【1019】 In a 1 L graduated cylinder, add 30 mL of 25x Wash Buffer Concentrate to 720 mL of MilliQ water. Mix until uniform. 【1020】 The wash buffer was stored at 4°C after preparation. 【1021】 5.2 Assay Buffer (LSbio Proprietary Ingredients) 【1022】 The sample diluents and assay diluents (assay diluents A and B) for the biotinylated antibody and neutravidin-HRP conjugate provided with the kit were used. 【1023】 5.3 Substrate 【1024】 The TMB substrate provided with the kit was used. 【1025】 5.4 Stop solution 【1026】 The stop solution provided with the kit was used. 【1027】 5.5 Detection Reagent A Working Solution 【1028】 Immediately before use: Dilute Detection Reagent A with Assay Diluent A at a ratio of 1:100. 【1029】 For example: If 11 mL of Detection Reagent A working solution needs to be prepared, add 110 μL of Detection Reagent A to 10,890 μL of Assay Diluent A. 【1030】 5.6 Detection Reagent B Working Solution 【1031】 Immediately before use: Dilute Detection Reagent B with Assay Diluent B at a ratio of 1:100. 【1032】 For example: If 11 mL of Detection Reagent B working solution needs to be prepared, add 110 μL of Detection Reagent B to 10,890 μL of Assay Diluent B. 【1033】 5.7 Assay Controls 【1034】 A control with an LPL level of 62.5 pg / mL, which falls within the range of the assay, is prepared. Single-use aliquots are prepared and stored at nominally -80°C. 【1035】 For example, use 200 μL of sample diluent to mix with 200 μL of LPL standard concentration 125 pg / mL; 【1036】 6 Preparation of standards and spikes 【1037】 6.1 Preparation of standard curve solutions 【1038】 6.1.1 For older lots (2019-2020), reconstitute the lyophilized standard using 2 mL of sample diluent to prepare an LPL stock concentrate at a concentration of 2000 pg / mL. Incubate the LPL stock at room temperature for 10 minutes without vortexing or vigorous mixing. 【1039】 6.1.2 For the latest lot of kit (from 2021), reconstitute the lyophilized standard with 1 mL of sample diluent to prepare an LPL stock concentration of 1000 pg / mL and incubate at room temperature for 10 minutes while vortexing or vigorously mixing. 6.1.3 Generate standard curve dilutions by serially diluting the LPL stock concentrate (2000 pg / mL) using sample diluent from step 6.1.1 to the following concentrations: 1000, 500, 250, 125, 62.5, and 31.25 pg / mL. See the table below for an example. 【1040】 [Table 48] 【1041】 6.1.4 Generate standard curve dilutions by serially diluting the 1000 pg / mL LPL stock concentrate from step 6.1.2 to the following concentrations: 500, 250, 125, 62.5, and 31.25 pg / mL using sample diluent. See the table below for an example. 【1042】 [Table 49] 【1043】 NOTE: Depending on the lot of commercial kit used for the assay, next use step 6.1.3 or 6.1.4 to dilute the standard curve at concentrations ranging from 1000 pg / mL, 500 pg / mL, 250 pg / mL, 125 pg / mL, 62.5 pg / mL, and 31.25 pg / mL. 【1044】 6.2 Preparation of assay spikes 【1045】 Prepare a high-level LPL assay spike of 250 pg / mL standard by diluting 1000 pg / mL (standard 1 from step 6.1) 4x with sample diluent. 【1046】 For example, dilute 200 μL of 1000 pg / mL (Standard 1) with 600 μL of Sample Diluent to a final concentration of 250 pg / mL. 【1047】 Prepare a mid-level LPL assay spike of 125 pg / mL by diluting 500 pg / mL (standard substance 2 from step 6.1) 4x with sample diluent. 【1048】 Prepare a low-level LPL assay spike of 31.25 pg / mL by diluting 125 pg / mL (standard substance 4 from step 6.1) 4x with sample diluent. 【1049】 Load triplicate spikes at 100 μL per well into a 96-well microtiter plate. 【1050】 6.3 Sample preparation 【1051】 6.3.1 Risankizumab Process 4 BDS, 150 mg / mL, is diluted to a concentration of 0.075 mg / mL for spiked sample preparation and the nominal dilution of the risankizumab BDS sample. 【1052】 [Table 50] 【1053】 6.4 Preparation of spiked samples 【1054】 6.4.1 In a polypropylene microtube, mix 200 µL of Abcam Sample Diluent with 200 µL of pre-diluted risankizumab BDS sample (step 6.3.1) in an equal volume to the unspiked risankizumab BDS sample. 【1055】 In a polypropylene microtube, mix 200 µL of 500 pg / mL standard solution (standard substance 2 from step 6.1) with 200 µL of pre-diluted risankizumab BDS sample (step 6.3.1) as a high-level PLA2 spiked sample. 【1056】 In a polypropylene microtube, mix 200 µL of 250 pg / mL standard solution (standard substance 3 from step 6.1) with 200 µL of pre-diluted risankizumab BDS sample (step 6.3.1) as an intermediate-level PLA2 spiked sample. 【1057】 In a polypropylene microtube, mix 200 µL of 125 pg / mL standard solution (standard substance 4 from step 6.1) with 200 µL of pre-diluted risankizumab BDS sample (step 6.3.1) as an intermediate-level PLA2 spiked sample. 【1058】 7. Procedure 【1059】 7.1 Assay Procedure: The checklist can be used as a guide by checking off completed steps. Reverse pipetting was used throughout unless otherwise stated. 【1060】 7.1.1 Allow all contents of the kit to stand at room temperature (RT) for at least 2 hours. 【1061】 7.1.2 Prepare standards, unspiked samples, spiked samples, and controls. 【1062】 7.1.3 Using a multichannel pipette, pipette 100 μL / well of standards, unspiked samples, spiked samples (if applicable), and controls into triplicate wells of the plate, and pipette 100 μL / well of assay buffer into all empty wells to serve as blanks. Cover with sealing tape and incubate at 37°C with shaking at approximately 100 rpm for 2 hours. 【1063】 7.1.4 Aspirate the liquid from each well and discard into a sink. Gently tap the plate on a paper towel or Kimwipe to remove any liquid residue; do not wash afterwards. 【1064】 7.1.5 Add 100 µL / well of Detection Reagent A working solution (step 5.5). Cover with sealing tape and incubate at 37 °C with shaking at approximately 100 rpm for 1 h. 【1065】 7.1.6 Aspirate the liquid from each well and discard into a sink. Gently tap the plate on a paper towel or Kimwipe to remove any residual liquid. Wash the plate three times with 350 μL / well of wash buffer. Allow the plate to incubate for 2 minutes for each wash, then aspirate the wash buffer. Pat the plate dry on a paper towel. 【1066】 7.1.7 Add 100 µL / well of Detection Reagent B working solution (step 5.6). Cover with sealing tape and incubate at 37 °C with shaking at approximately 100 rpm for 1 h. 【1067】 7.1.8 Aspirate the liquid from each well and discard into a sink. Gently tap the plate on a paper towel or Kimwipe to remove any residual liquid. Wash the plate five times with 350 μL / well of wash buffer in each well. Allow the plate to incubate for 2 minutes for each wash, then aspirate the wash buffer. Pat the plate dry on a paper towel. 【1068】 7.1.9 Add 90 µL / well of TMB substrate (step 5.3), cover with sealing tape, protect the plate from light, and incubate at 37 °C for 10 min without shaking (start timer as soon as you add the substrate to the first column). 【1069】 7.1.10 Stop the reaction by adding 50 µL / well of stop solution (step 5.4). 【1070】 7.1.11 Read plate at 450 nm within 2 minutes after adding stop solution. Blank subtraction was performed. 【1071】 8 Data analysis and calculations 【1072】 NOTE: Only samples and control dilutions with OD values within the 31.25 pg / mL standard (assay LOQ) and 1000 pg / mL (highest standard) and passing the spike recovery criteria (if applicable) were accepted. If a sample is below the 31.25 pg / mL standard (less than 31.25 pg / mL) or LOQ, the result should be reported as less than 31.25 pg / mL or less than LOQ. This result should then be multiplied by the dilution factor and divided by the initial sample concentration (mg / mL) to report the result in pg / mg. If the sample had a high LPL concentration and the sample exceeded the standard curve (1000 pg / mL), it was repeated at a dilution high enough to fall within the standard curve. If the sample could not be spiked, repeat the dilution. 【1073】 [Table 51] 【1074】 8.1 Standard curve 【1075】 8.1.1 Standard concentrations must be entered into the protocol template. 【1076】 8.1.2 Graph the standard curve using each response value (OD) using 4-parameter curve fitting (4P). 【1077】 Note: The above 4-parameter fitting equation is equivalent to: 【number】 【1078】 8.1.3 If the reference material does not meet the acceptance criteria, the assay should be repeated. 【1079】 8.2 Spiked Samples (if applicable) 【1080】 8.2.1 For spiked results (assessed per well) that were within the assay range: Multiply the spiked sample results by the dilution factor. Calculate the mean and %CV of the dilution-corrected spiked sample results. The %CV should be ≤ 20%. Calculate the % spike recovery using the formula below: Spike Recovery % = ((Spiked Sample Result (pg / mL) - Unspiked Sample Result (pg / mL)) ÷ (Theoretical Spike (S) (pg / mL)) x 100 【1081】 8.2.2 The recovery of spikes based on average results must be within 50% to 150%. 【1082】 8.2.3 If this criterion is not met, the sample must be repeated. 【1083】 8.3 Unspiked Samples 【1084】 8.3.1 For sample dilutions within the assay range (assessed per well): Calculate the mean and %CV of the dilution-corrected results obtained. The %CV between dilutions should be ≤20%. If this criterion is not met, the sample should be repeated. 【1085】 8.3.2 Using the acceptable results, calculate the LPL concentration (ng / mg) by dividing the average result, corrected for the final dilution factor, by the initial sample stock concentration. 【1086】 【number】 【1087】 8.3.3 Record the final result to a whole number. 【1088】 8.4 Contrast 【1089】 8.4.1 For control dilutions within the assay range (assessed per well): Calculate the mean and %CV of the dilution-corrected results obtained. The %CV between dilutions should be ≤ 20%. If this criterion is not met, the assay should be repeated. Record the final result in ng / mL to one decimal place. If the control is outside the specified range, the assay should be repeated. 【1090】 8.5 Assay range: The assay range was 31.25 pg / mL to 1000 pg / mL. 【1091】 result 【1092】 The PLA2, PLBL2, and LPL levels in various lots of DP1, DP2, DP3, and DP4 were determined and are shown in Table 47 below. 【1093】 [Table 52] 【1094】 As shown in Table 47, the levels of PLBL2 are significantly reduced in DP3 and DP4 compared to DP1 and DP2. For example, in DP4, PLBL2 levels are reduced by almost 1000-fold compared to DP1 and DP2. 【1095】 Similarly, the levels of PLA2 are significantly reduced in DP3 and DP4 compared to DP1 and DP2, which have at least 0.25 ng / mg (250 pg / mg) of PLA2. 【1096】 Example 10: Polysorbate stability is increased in risankizumab drug products DP3 and DP4 The stability of polysorbate 20 (PS20) and polysorbate 80 (PS80) was determined by detecting the levels of PS20 or PS80 and free fatty acids (FFA) in samples stored at various temperatures (i.e., 5°C, 25°C, and 40°C) for 6 months. 【1097】 Determination of PS20 / PS80 using a charged aerosol detector (CAD) 【1098】 The method for detecting the level of PS20 in various pharmaceuticals is described below. The level of PS80 in various pharmaceuticals was determined using the same method. 【1099】 1. Principles 【1100】 The polysorbate 20 content in 150 mg / mL formulation test samples was determined using a charged aerosol detector (CAD). Detection was based on nebulization of the analyte in a continuous nitrogen stream. After removal of the mobile phase, a second positively charged nitrogen stream formed charged particles. The released charge was proportional to the amount of polysorbate 20. 【1101】 2. Equipment 【1102】 An HPLC system equipped with gradient elution, a temperature-controlled autosampler, a column oven, and a degassing unit, as well as stainless steel capillaries (for the HPLC column and for connecting the column to the detector). 【1103】 Corona Veo RS Charged Aerosol Detector (CAD), Thermo Fisher Scientific part number 5081.0020 【1104】 Chromatography data system (e.g., Empower) 【1105】 Column: Waters Oasis MAX column (2.1 x 20 mm, 30 μm), part number 186002052 【1106】 HPLC Amber Vials, Screw Cap, Agilent, Cat. No. 5182-0716 or equivalent 【1107】 Blue screw cap, Agilent, catalog number 5182-0717 or equivalent 【1108】 Analytical balance device 【1109】 pH meter 【1110】 Stir plates, stir bars, and vortex mixers 【1111】 volumetric flask 【1112】 3.Material 【1113】 Ammonium formate, Fisher, LC-MS grade, catalog number A115-50 or equivalent 【1114】 Methanol, EMD, LC-MS grade, catalog number MX0486-6 or equivalent 【1115】 Isopropyl alcohol, Fisher, LC-MS grade, catalog number A461-212 or equivalent 【1116】 Acetonitrile, Fisher, Optima LC-MS grade, catalog number A955-212 or equivalent 【1117】 Formic acid, ThermoScientific, catalog number 28905 or equivalent 【1118】 Polysorbate 20 (PS20), JT Baker, catalog number 4116-02 or equivalent 【1119】 Project GB Reference Standards 【1120】 Purified water (Type 1 grade, e.g., Milli-Q, WFI, or equivalent) 【1121】 4. Preparation of Solutions 【1122】 4.1 Dilute formic acid solution (1:1) for adjusting the pH of mobile phase A 【1123】 4.1.1 Add 7.5 mL of formic acid and 7.5 mL of purified water to a glass beaker. 【1124】 4.1.2 Mix the solution until homogeneous. 【1125】 4.2 Mobile phase A (10 mM ammonium formate pH 3.0 / 20% isopropyl alcohol) 【1126】 4.2.1 Add 1400 mL of purified water to a 2 L beaker. 【1127】 4.2.2 Weigh out 1.26g ± 0.01g of ammonium formate and add to the beaker. 【1128】 4.2.3 Adjust the pH of the solution to 3.0 ± 0.1 using the diluted formic acid solution from step 4.1. 【1129】 4.2.4 Add 400 mL of isopropyl alcohol. 【1130】 4.2.5 Transfer the solution to a 2 L volumetric flask. 【1131】 4.2.6 Add purified water to make the volume the same. 【1132】 4.2.7 Transfer the solution to a suitable container. 【1133】 4.2.8 Mix the solution for approximately 15 minutes. 【1134】 4.2.9 Check the homogeneity of the solution by visual inspection. 【1135】 4.2.10 Store at room temperature for up to one week. 【1136】 4.3 Mobile phase B (50% isopropyl alcohol / 50% acetonitrile) 【1137】 4.3.1 Add 500 mL of isopropyl alcohol and 500 mL of acetonitrile to a 1 L glass bottle. 【1138】 4.3.2 Mix the solution for approximately 15 minutes. 【1139】 4.3.3 Check the homogeneity of the solution by visual inspection. 【1140】 4.3.4 Store at room temperature for up to one month. 【1141】 4.4 Autosampler Rinse Solution (20% Methanol) 【1142】 4.4.1 Add 200 mL of methanol and 800 mL of purified water to a 1 L glass bottle. 【1143】 4.4.2 Mix the solution until homogeneous. 【1144】 4.4.3 Store at room temperature for up to one week. 【1145】 5. Preparation of Standards, Samples, and Blanks 【1146】 5.1 Recommendations for the generation of standards and samples 【1147】 5.1.1 Accurate Pipetting of Polysorbate 20 【1148】 Care should be taken when pipetting PS20 containing samples, standards, and diluents. It is recommended to equilibrate the pipette tip by aspirating and dispensing five times. Pipetting should be done slowly. It is important to wait for viscous solutions to be completely expelled and, after aspirating, for the thin film of solution to completely leave the pipette tip. When dispensing, it is equally important to wait for the solution to completely disappear from the pipette tip. Waiting approximately 15 seconds in each direction (aspirate / dispense) may be necessary. 【1149】 5.1.2 Volume of standard solution and sample solution 【1150】 The volumes of the calibration standards and diluted sample solutions may deviate from 500 μL, provided that the volumes of the standards and diluted sample solutions are the same. 【1151】 5.2 10 mg / mL Polysorbate 20 Stock Solution I 【1152】 5.2.1 Weigh out 1.00 g ± 0.01 g of polysorbate 20 into a tared 100 mL volumetric flask. 【1153】 5.2.2 Add purified water to make the volume. 【1154】 5.2.3 Carefully add a stir bar and appropriate cap. If the flask is not light-tight, wrap the flask in foil. 【1155】 5.2.4 Stir and mix for approximately 15 minutes. 【1156】 5.2.5 Check the homogeneity of the solution by visual inspection. 【1157】 5.2.6 Store at 2-8°C for a maximum of 1 week. 【1158】 5.3 0.5 mg / mL Polysorbate 20 Stock Solution II 【1159】 5.3.1 Before preparation, stir Polysorbate 20 Stock Solution I for at least 10 minutes at room temperature. This is only required if Solution I is not freshly prepared. 【1160】 5.3.2 Dilute 2.5 mL of Polysorbate Stock Solution I into a 50 mL volumetric flask. 【1161】 5.3.3 Add purified water to make the volume. 【1162】 5.3.4 Stir to mix for approximately 15 minutes. If the flask is not light-tight, wrap it in foil. 【1163】 5.3.5 Check the homogeneity of the solution by visual inspection. 【1164】 5.3.6 Store at 2-8°C for a maximum of 1 week. 【1165】 5.4 Calibration solutions 【1166】 For the calibration curve, prepare a minimum of six calibration standards with different PS20 concentrations using Polysorbate 20 Stock Solution II from Section 5.3. An exemplary dilution scheme is shown below in Table 48. All dilutions are prepared in water. Volumes may be adjusted proportionally, ensuring the target PS20 concentration remains unchanged and the total volume of each preparation is greater than 10 mL. 【1167】 5.4.1 Prepare a polysorbate 20 standard curve from 0.0125 mg / mL to 0.15 mg / mL. Add purified water to the volumetric flasks to make the volumes equal as follows: 【1168】 [Table 53] 【1169】 5.4.2 Mix by inverting approximately 15 times. If the flask is not light-tight, wrap it in foil. 【1170】 5.4.3 Store at 2-8°C for a maximum of 1...

Claims

[Claim 1] A liquid composition comprising (1) risankizumab and (2) phospholipase A2 (PLA2) in an amount of less than approximately 250 pg per 1 mg of risankizumab.