They reported a reduction in tumor uptake of 41%. TGF-beta Immunohistochemistry confirmed the decrease in Her2 expression ex vivo in a qualitative way only. KramerMarek and colleagues measured changes in Her2 expression after 17DMAG treatment with the same affibody as we used in our study, but instead of using optical imaging they labeled the affibody with fluorine18 for PET imaging and only carried out a single pre and posttreatment scan . They reported a reduction of33% in an MCF7 cell line transfected with Her2 and of 71% in BT474 breast tumor xenografts. Her2 downregulation was confirmed ex vivo by Western blot and ELISA. Both Oude Munnink and colleagues and KamerMarek and colleagues compared a single posttreatment measurement with pretreatment . Her2 expression was not monitored over a longer period of time.
The strength of our study is that we followed each mouse over 10 days, which enabled us to see the Her2 levels decrease Fluorouracil after treatment and recover after the treatment was stopped . This indicates that we can monitor the molecular changes noninvasively over time with our optical imaging strategy, whereas we did not observe significant changes in tumor volume during the study. Our in vivo results of 22.5% signal reduction are consistent with the previous reports, considering that different cell lines were used for the tumor xenografts and that the imaging technique used was also different. In addition, correlating in vivo optical imaging signal with ex vivo Her2 levels by Western blotting further supported our results.
Although tumor volume did not change significantly after 17 DMAG treatment, in 2 of the 17DMAG–treated mice the clone B tumors shrunk to very small volumes at day 9. To confirm that the changes in optical imaging signal were due to a decrease cell nucleus in Her2 expression levels and not caused by other nonspecific antitumor effects of the drug, we correlated the in vivo optical imaging signal with the ex vivo Her2 expression levels not only at day 9, but also at day 3 in a subgroup of 8 mice. In that group, we also closely monitored the tumor volume by ultrasound measurements up to day 3 and confirmed that there was no decrease in tumor volume after treatment . Results indicate that the measured changes in optical imaging signal reflect the changes in Her2 expression after drug treatment.
An important advantage of optical imaging in comparison with PET imaging is that it does not use radioactive components or ionizing radiation and can thus be used more frequently. An additional advantage is that optical imaging agents are easier to generate and much less expensive than PET tracers. In contrast to radionuclide imaging approaches in which over time the imaging signal disappears as a result of natural decay in addition to clearance from the subject, in optical imaging the clearance of the imaging signal is predominantly dependent on clearance of the imaging probe from the body. For this reason, small molecules with quick clearance, such as affibody molecules, may be preferable over large molecules in optical imaging. In addition, preinjection optical signal can be measured and subtracted from subsequent imaging examinations in optical imaging to adjust for the residual signal as done in this work.