Methods
The nanobioconjugate (a potent anticancer drug candidate), along with the drug vehicle (PMLA), positive (PHA-M) and negative (PBS) controls, was tested for immunogenicity (cytokine release). Nanodrug was added in vitro to the whole blood samples from three healthy donors at therapeutic 1X dosage (based on the expected Cmax in human at the human equivalent to the mouse efficacy dose), or 3X dosage. Diluted blood was incubated with controls or nanomaterial for 24 hours. Using quantitative Meso Scale Discovery (MSD) assay, nine cytokines (TNFa, IFNgamma, IL12p70, IL1b, IL2, IL4, IL6, IL8, and IL10) in the donors' blood were simultaneously measured.
Results
A therapeutic 1X concentration and even 3X elevated concentration of the nanodrug did not significantly induce any of the nine cytokines in the donors' blood except for IL8 in the 3X elevated dose group. However, this IL8 level was still 40-fold lower than in the positive control group.
Conclusion
These data showed that there is potentially low risk of the nanodrug containing antibodies and oligos to induce severe immunogenic response (“cytokine storm”).
Other immunogenic in vitro tests (e.g., T4 lymphocyte immunogenic response in PBMC) are also available for testing of the new therapeutic proteins and other biologics.