Table 1: CE-MS: Unexploited Opportunities in Process Development?

Transcription

1 Table 1: CE-MS: Unexploited Opportunities in Process Development? Session 1 Facilitator: Mei Han, Amgen Inc. Scribe: Yunan Wang, Amgen Inc. Session 2 Facilitator: Göran Hübner, Boehringer Ingelheim Pharma GmbH & Co. KG Scribe: Tara Enda, Bristol-Myers Squibb Company Scope: Capillary Electrophoresis-Mass Spectrometry (CE-MS) has become an increasingly important technique over the last two decades for biopharmaceutical industry with the improvement of the CE-MS interface and MS instrumentation. CE-MS is an orthogonal technique and provide complementary information to LC-MS. For example, CE-MS can provide intact and bottom up protein characterization, charge variant identification, glycoprotein profiling, protein-ligand complex, protein integrity in vitro and in vivo, as well as in biomarker characterization. This table will focus on the Unexploited Opportunities in Process Development discussions and explore the future directions. Questions for Discussion: 1. What types of CE-MS methods do you think are promising to add value to cqa monitoring during process development, compared to the existing and commonly used method panel? (like e.g. CE-MS- based glycan profiling, CE-MS-based peptide mapping, or CE-MS-based top-down / middle-down approaches) 2. What are your ideas to ensure high throughput commonly requested from process development departments? 3. What are your ideas (or is your experience) to keep (relative) quantification of cqa in CE-MS methods at reasonable intermediate precision levels, especially with respect to multiple MS instruments applied, or with respect to (long) regular development periods? 4. Would CE-MS during process development represent an option for a MAM approach? 5. What are barriers for implementing CE-MS (including chip-ms) in development of biopharmaceuticals? What s the advantages and challenges for implementing CE-MS? 6. What are the existing techniques and how does CE-MS to help with definitively identify all the peaks in CE separations? 7. Which new applications could be of high interest during protein characterization and process development?

2 Discussion Notes: Session 1: Common CE-MS systems on the market o CMP Scientific EMASS II system Nano-flow sheath liquid CE-MS interface, based on Dr. Dovichi s design o 908 Devices ZipChip Chip-based CE-MS interface, based on Dr. Ramsey s design o SCIEX CESI 8000 plus Sheathless CE-MS interface, based on Dr. Moini s design o Agilent Sheath liquid CE-MS interface Distinguished advantages of CE-MS o CE-MS is compatible with small sample volume. o No run to run carryover o Salts and matrices could be separated from the target analytes. o CE has small reagents consumption. Applications of CE-MS in biotech and pharmaceutical industries o In the product development and production functions: They usually have a larger amount of the material. ZipChip can be a simple and fast analytical method in these areas. o In the pharmacokinetics function: In vivo samples, for example, serum, plasma, urine, saliva etc, usually have lower concentrations and limited sample volumes. The capillarybased CE-MS systems fit better for these studies. o CIEF-MS Characterization of charge heterogeneity BMS collaborated with CMP and published a paper about on-line CIEF- MS Anal. Chem., 2018, 90 (3), pp Advanced Electrophoresis Solutions Ltd. has the icief fractionation technology. Each fractionation can be further analyzed by MS off-line. CZE could be more compatible with MS, but it might have the best resolution. Sample preparation challenges o The CE-MS instrument is getting more and more robust, the key to achieve the best data quality in the complicated sample matrix is the sample preparation. o Better sample prep=better signal o Immunoaffinity purification Magnetic beads based immunoaffinity purification Analytical Biochemistry 2017, 539: Agilent AssayMap system Faster, more reproducible, cleaner eluted samples

3 Data analysis challenges o Similar with LC-MS, the difficulty of the mass spec data interpretation depends on the experience level of the analyst. o The software from each vendor has both pros and cons. o Protein matrix, as a vendor-neutral software, might have a good intact mass analysis algorithm How do we make CE-MS a more routine workflow? o Try to make the platform method that fit for multiple constructs with a single method. o Try to train people without any CE experience at all to use CE-MS. How about reagent kits? o Good for QC and CRO o CE use so little buffer. It probably is not a good business for the vendors. o The reagent composition of the kit usually becomes proprietary. It might not be good for method development. Wishlist for CE-MS vendors o Plate format autosampler o Improve the robustness of the system o Develop new capillary coatings o Vendor-neutral capillaries/cartridges Session 2: What types of CE-MS methods do you think are promising to add value to cqa monitoring during process development, compared to the existing and commonly used method panel? What is most desired application for CE-MS? (Answers are listed in no particular order) o CE-MS may provide answers to unknowns / things that may have been missed or excluded by HPLC-MS. For example small peptides that are not retained on peptide map columns may be seen by CE-MS. Another example Host cell protein (HCP) characterization- assess if CE- MS can gather equivalent or additional information in comparison to LC- MS data. o Charge heterogeneity profile is highly desired but question on which approach is practical was noted: Classical CE methods with cleanup step to enable CE-MS MS friendly charge heterogeneity method o Size based Any advantages to size based intact CE-MS vs size based intact LC-MS? o Peak characterization - might enable faster turnaround time than collecting fractions from LC method Charge based qc method peak characterization Size based qc method peak characterization o Other: Hydrophobicity Methods Glycan analysis

4 o Look towards to the future: more complex molecules current charge heterogeneity methods may not be feasible and may require mass spec analysis. Opportunity for CE-MS. What are your ideas to ensure high throughput commonly requested from process development departments? o Process development samples run via CE-UV type method with ability to connect to Mass Spec for MS peak characterization. o Push for investment in new methods. o Automation: CE-MS will require this!!!!! If I need to now assess process development samples and formulation study samples...there will a high amount of samples... automation on all ends of the CE-MS connections will be necessary. What are your ideas (or is your experience) to keep (relative) quantification of cqa in CE-MS methods at reasonable intermediate precision levels, especially with respect to multiple MS instruments applied, or with respect to (long) regular development periods? o Critical quality attribute testing are being requested at earlier and earlier phases of product development. Early Process Development samples typically are not high in physical volume and are at lower sample concentrations than later phased products. Sensitive CE-MS method(s) that accommodate low sample quantity and low concentration would be needed for early process development samples. What are barriers for implementing CE-MS (including chip-ms) in development of biopharmaceuticals? What s the advantages and challenges for implementing CE-MS? o Mass Spec Interference Understanding Need to understand more from Mass Spectrometer point of view what materials interfere in Mass Spec analysis. Limit one run and then clean Mass Spec? What reagents interfere in Mass Spec and at what quantities? Example small amount of ampholytes gets into Mass Spec correlating to more Mass Spec maintenance. Is this an issue? o Shealth flow vs no Shealth flow might need to use shealth flow to enable better cleanup before mass spec repeatability vs sensitivity o Software Mass Spec software is good as is and anticipate incorporation of CE into MS would include incorporation of data into current MS software programs / data analysis platforms. Missing perhaps may be mitigation of the connection readouts for voltage from CE or microchip component which assume would be with the CE or CE interface itself. o Advantage CE-MS Physics ion suppression = high flow vs low flow = physics = reliable low flow should = lowest ion suppression best case for MS - signal = true value and not an ion suppressed estimation.

5 Mass Spec itself should not be challenge unless you are CE expert and not Mass Spec expert. Then you need to hire a Mass Spec expert. cief method development is the challenging part, MS data analysis part is not the challenge once CE and MS are paired together. o Separation bias with regards to resolution may occur / CE method design question. For example resolution of basic peaks might be different from acidic peaks and vice versa depending on CE-MS separation approach taken and how this will be incorporated into CE-Mass Spec method development. Extrapolate the differences in CE and LC to get full resolution CE-MS o Robustness CE-MS methods would need to be completely robust. Robustness is big factor. Can t be one run and done. Can t work one day and then down for next week. Needs to be quick assembly / easy to use and start up assay for success. Would CE-MS during process development represent an option for a MAM approach? o CE MS MAM (multi-attribute method) technology. What does MAM do that requires LC? Why can t MAM be CE-MS MAM platform? Which new applications could be of high interest during protein characterization and process development? o Wave of the future for process impurities and product degradation identification and quantification. o Why stop with production...why not include clinical aspect and truly harness the ability to concentrate low volume low concentration samples