Biotherapeutics are a promising class of molecules in drug discovery, but they are often limited to extracellular targets due to their poor cell penetration. High-throughput cell penetration assays are required for the optimization of biotherapeutics for enhanced cell penetration.
Researchers in the Kritzer Lab, have developed a HaloTag-based assay called the chloroalkane penetration assay, CAPA, which is quantitative, high-throughput, and compartment-specific. The lab members, published in JACS, demonstrate the ability of CAPA to profile extent of cytosolic penetration with respect to concentration, presence of serum, temperature, and time.
They also used CAPA to investigate structure–penetration relationships for bioactive stapled peptides and peptides fused to cell-penetrating sequences. CAPA is not only limited to measuring cytosolic penetration. Using a cell line where HaloTag is localized to the nucleus, the lab members show quantitative measurement of nuclear penetration. Going forward, CAPA will be a valuable method for measuring and optimizing the cell penetration of biotherapeutics.
Cell Penetration Profiling Using the Chloroalkane Penetration Assay
Leila Peraro, Kirsten L. Deprey, Matthew K. Moser, Zhongju Zou, Haydn L. Ball, Beth Levine, and Joshua A. Kritzer
J. Am. Chem. Soc., 2018, 140 (36), pp 11360-11369