Stability of Recombinant Human Thyrotropin Potency Based on Bioassay in FRTL-5 Cells

Abstract

Background:

Recombinant human thyrotropin (rhTSH; Thyrogen^®) is approved for use in a 0.9 mg dose/day for 2 consecutive days for diagnosis and treatment of differentiated thyroid cancer. It is recommended that it be injected immediately after reconstitution in the distilled water diluent supplied by the manufacturer. However, Thyrogen has been used off-label in doses less than the standard 0.9 mg dose for stimulation of radioiodine uptake in the treatment of multinodular goiter. To determine whether the biologic activity of Thyrogen can be preserved after dilution, we designed experiments to assess the biologic stability of Thyrogen under different durations and temperatures of storage.

Methods:

rhTSH was diluted in 1% bovine serum albumin in phosphate-buffered saline to a concentration of 0.9 mg/mL and further diluted to 0.1 mg/mL. Aliquots of 0.5 mL were stored at room temperature, 4°C, −11°C, and −60°C for various lengths of time. In addition, rhTSH aliquots were also subjected to incubation for 1 hour at 50°C and to 10 cycles of freezing in dry ice alternating with thawing at 37°C. Bioassays were performed in FRTL-5 cells. rhTSH was added to the media at a final concentration of either 5 ng/mL or 20 ng/mL, and the cells were then incubated for 48 hours. Potency was assessed by measurement of ¹²⁵I-iodide uptake in comparison to cells treated with perchlorate to block iodide uptake. Samples were immunoassayed at day 185 of storage.

Results:

Samples stored at 4°C, −11°C, −60°C, and room temperature retained activity after storage periods of up to 204 days. Samples subjected to 10 freeze–thaw cycles or heated to 50°C for 1 hour retained full biologic activity. Immunoassay at day 185 showed no difference in immunoactivity in relation to the storage condition.

Conclusion:

rhTSH kept at 4°C, −11°C, −60°C, and room temperature maintained good biologic potency for more than 6 months of storage when tested in vitro, indicating that the biologic activity is very stable. However, altered sialylation occurring during storage could have altered the half-life of rhTSH. Nevertheless, the data provide reassurance that storage in the cold for a few months does not result in significant loss of biologic activity.