When formulating protein products, it is vital that proteins react with excipients rather than themselves. Eric Munson, Professor and Head, Purdue University, discussed stability prediction for lyophilised formulations. He suggested that solid-state NMR spectroscopy is a critical analytical technique that is highly useful for predicting the stability of protein formulations.
Water content and a high level of mobility are common issues in protein formulations, but adding bulking agents like mannitol, sucrose, and trehalose can overcome these issues. Munson also said that buffers like histidine and phosphate contribute to stabilising protein formulation by preventing protein aggregation and maintaining ionisation states.
There are already many different analytical techniques available for characterising proteins, so what gives solid-state NMR spectroscopy an edge? NMR spectroscopy allows scientists to observe different crystalline forms that may be present in a system. It can also measure proton relaxation times, which gives insights into mobility and miscibility. Overall, NMR is the most effective at measuring the fraction of crystalline vs amorphous.
Munson also introduced the use of time-domain NMR to measure water content on lyophilised samples. He reiterated its ability to distinguish between rigid and mobile water components and the wider impact of this on protein stability. For instance, after secondary drying, mobility drops significantly, indicating better stability. Solid-state NMR can determine whether water is locked in the matrix or free to move.
Next, Munson presented some novel findings on the miscibility of proteins with excipients at various nanometre scales. Studies involving IgG and LDH proteins, as well as excipients such as trehalose and dextran, demonstrate that trehalose generally supports homogeneity and stability, while dextran typically leads to phase separation and instability.
Real-world studies posit that sucrose is slightly more stable than trehalose because trehalose crystallises under specific conditions, potentially leading to phase separation. However, Munson still implied that trehalose is still a good contender for stability. The study also examined mannitol, which is prone to crystalllisation, therefore diminishing its effectiveness as a stabiliser when used alone.
Munson concluded by outlining his future plans are to explore combinations like mannitol with trehalose or sucrose and using C13-labelled compounds for enhanced resolution. He also reaffirmed that his goal is to define optimal excipient ratios for maximum protein stability.
