RNA-based therapeutics, including siRNA and mRNA, hold significant promise in the pharmaceutical landscape. Several RNA-based therapeutics have been granted FDA approval for vaccines and treating genetic diseases. Lipid nanoparticles (LNPs), play a crucial role in overcoming biological barriers during siRNA and mRNA delivery. The efficacy and safety of RNA-loaded nanoparticles are influenced by critical characteristics such as particle size, shape, surface charge, surface area, ionization constant, and aggregation (Patel).
The pKa value of ionizable lipids, used in LNPs, significantly influences the success of RNA delivery into cells. The apparent pKa is the pH where the quantities of ionized and deionized groups within the LNP are balanced within the systems. It governs the ionization behavior and surface charge. This parameter significantly impacts the stability, potency, and toxicity of these nanoparticles. The apparent pKa serves as a gauge for estimating the surface charge and ionic interactions within assembled nanomaterials in nanoparticles, so optimizing the apparent pKa of nanoparticles is essential for effective and safe RNA formulations. The best performing lipid and polymer nanoparticles for RNA delivery exhibit apparent pKa values between 6 and 7 (Patel). LNPs with pKa values equal to or less than 5.4 do not ensure in vivo efficacy and can be ruled out (Whitehead).
One notable ionizable lipid applied in FDA-approved drugs Onpattro and Givlaari for the delivery of RNA is D-Lin-MC3-DMA (Figure 1), which has a pKa of 6.44 (Patel).
Figure 1. Ionizable Lipid D-Lin-MC3-DMA (pKa 6.44)
For maximizing the potency of hepatic delivery of siRNAs, LNPs with an optimized pKa in the range of 6.2 to 6.4 prove effective. Within this range, LNPs strike the right balance of charge and stability. L319 (Figure 2) is an MC3 analog with a pKa in this range (Patel). In vivo, L319 demonstrates superior delivery efficacy and quicker elimination from the liver and plasma compared to MC3 (Hou).
Figure 2. Ionizable Lipid L319 (pKa 6.38)
The pKa value also influences the immunogenicity of LNPs, with an optimal range of 6.6 to 6.9 for intramuscular (IM) delivery. Lipids within this range are chosen for mRNA vaccine delivery due to their strong biodegradability, tolerability, protein expression capabilities, and favorable immunogenicity (Patel). Among the significant lipids, SM-102 (Figure 3) stands out, playing a crucial role in the formulation of the FDA-approved, COVID-19 vaccine mRNA-1273. SM-102 exhibits a low immunogenicity, rapid clearance rate, and improved safety profile.
Figure 3. Ionizable Lipid SM-102 (pKa 6.68)
One subclass of ionizable lipids garnering recognition and contributing considerably to LNP formulations is multi chargeable lipids. They are exemplified by having more than two hydrophobic tails, like C12-200 (Figure 4), which is commonly employed as a benchmark in the quest for novel ionizable lipids. 304O13 (Figure 5) is a multi chargeable lipid with similar potency to C12-200. Moreover, it has the benefit of having lower toxicity at higher doses because of the presence of ester groups (Han), and it has a more favorable pKa than that of C12-200.
Figure 4. Multi chargeable Lipid C12-200 (pKa 6.96)
Figure 5. Multi Chargeable Lipid 304O13 (pKa 6.8)
In conclusion, an optimum pKa range of 6 to 7 is generally considered ideal for developing RNA-LNPs (Patel).
BroadPharm offers a wide variety of ionizable and other lipids to further fine-tune the pKa and other properties of RNA-LNP formulations. BroadPharm also offers fast-paced custom lipid syntheses to satisfy your research needs.
Patel, P., Ibrahim, N. M., & Cheng, K. (2021). The importance of apparent pKa in the development of nanoparticles encapsulating siRNA and mRNA. Trends in pharmacological sciences, 42(6), 448-460.
Whitehead, K. A., Dorkin, J. R., Vegas, A. J., Chang, P. H., Veiseh, O., Matthews, J., ... & Anderson, D. G. (2014). Degradable lipid nanoparticles with predictable in vivo siRNA delivery activity. Nature communications, 5(1), 4277.
Han, X., Zhang, H., Butowska, K., Swingle, K. L., Alameh, M. G., Weissman, D., & Mitchell, M. J. (2021). An ionizable lipid toolbox for RNA delivery. Nature communications, 12(1), 7233.
Hou, X., Zaks, T., Langer, R. et al. Lipid nanoparticles for mRNA delivery. Nat Rev Mater 6, 1078-1094 (2021). https://doi.org/10.1038/s41578-021-00358-0