The pharmaceutical stability of trastuzumab after short-term storage at room temperature assessed by analytical...

We report the pharmaceutical stability of trastuzumab stored for a short time (12 h) at room temperature (RT; 20-25 °C) compared to trastuzumab stored at 2-8 °C. The physicochemical properties were evaluated by UV-visible and FTIR spectroscopy, SDS-PAGE and size-exclusion HPLC (SE-HPLC). Trastuzumab was reacted with benzylisothiocyanate diethylenetriaminepentaacetic acid (BzDTPA) to complex ¹¹¹In. The HER2-binding affinity of ¹¹¹In-BzDTPA-trastuzumab synthesised from trastuzumab stored at RT or at 2-8 °C was measured using HER2-positive SK-Br-3 human breast cancer (BC) cells. The tumour and normal tissue uptake of ¹¹¹In-BzDTPA-trastuzumab was studied by microSPECT/CT imaging and biodistribution studies in CD1 athymic mice with s.c. HER2-positive SK-Ov-3 human ovarian cancer xenografts. There were no differences in λ_max or molar absorptivity (ε) values in the UV-visible spectra of trastuzumab stored at RT or at 2-8 °C. FTIR spectroscopy suggested no differences in secondary structure. SDS-PAGE revealed protein bands corresponding to the expected molecular weights. SE-HPLC showed identical properties for trastuzumab stored at RT or at 2-8 °C. The dissociation constant (K_d) for binding of ¹¹¹In-BzDTPA-trastuzumab to HER2 on SK-Br-3 cells (2.2-4.4 nM) was not significantly different when the radioimmunoconjugates were synthesised from trastuzumab stored at RT or at 2-8 °C. MicroSPECT/CT demonstrated high uptake in SK-Ov-3 tumours in mice that was not significantly different using trastuzumab stored at RT or at 2-8 °C (33.7 ± 8.8% vs. 22.2 ± 8.1% i.d./g, respectively; P = 0.36). There were no significant differences in normal tissue uptake or in tumour/normal tissue (T/NT) ratios. We conclude that short-term storage of trastuzumab at RT for 12 h did not affect the physicochemical or biological properties of the drug.