Application Notes

1 Formulation Development Application Note NT-PR-013 Use of iFormulate™ and nanoDSF for Fast and Precise Protein Formulation Development Dennis Breitsprecher*, Rajiv Nayar # , *NanoTemper Technologies GmbH, Floessergasse 4, 81369 Munich # HTD Biosystems, 1061 Serpentine Lane, Ste E, Pleasanton, CA 94566, USA Abstract One of the challenges in protein formulation is the simultaneous evaluation of multiple key formulation variables in a rational fashion. The Design of Experiments (DOE) approach has gained significant popularity in protein formulation as well as other process development activities. HTD Biosystems has developed a robust DOE approach for protein formulations referred to as iFormulate™ that utilizes advanced response surface quadratic modeling to evaluate the effects of four formulation variables in a multivariate fashion. The procedure allows for rapid identification of stable protein formulations, using only small amounts of material. By analyzing the thermal stability of a model protein, Lysozyme, using the Prometheus NT.48 in conjunction with the iFormulate TM DOE approach, we demonstrate that ideal formulations can be identified in as little as 35 minutes with minimal sample consumption. The DOE analysis of the precise and reproducible data generated by NanoTemper Technologies Prometheus NT.48 from 25 trial formulations resulted in the identification of the formulation design space for lysozyme formulations with high melting temperatures (T m s). The formulations can be rationalized using Quality by Design principles from the DOE analysis. Thus, by using the predictions from the DOE output and high-precision thermal stability analysis by nanoDSF, stable formulations of proteins can be generated and validated with unprecedented speed and precision. Introduction The new reality in biopharmaceutical drug development is that over the past 25 years, since the genome project, the pace of research has increased exponentially but the pace of development of new biopharmaceuticals has lagged behind. As a result, development groups are constantly challenged to bring an ever-growing number of biologics through the pharmaceutical development pipeline. Not only have the number of development projects increased dramatically, but the groups are under immense pressure to deliver the drug candidates under tight timelines, with limited material, and limited resources. Unfortunately, the billions of dollars spent in research have not translated into appropriate resources being dedicated to the development groups. The advent of three revolutions going on at the same time in the 21 st century such as the medical revolution, the computer revolution, and the quantum revolution has offered us the opportunity to address the challenges of the drug development pipelines. These revolutions have resulted in high throughput instrumentation that can generate an immense volume of data in a short period of time using

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