Figure 1.
Supposed structure of [99mTc]Tc(Tricin)nHYNIC-DAPI (99mTc-HYNIC-DAPI) with HYNIC as monodentate ligand.
Table 1.
Emission Spectrum of 99mTc (Data from Howell [17]).
Figure 2.
RP-HPLC Chromatogram of 99mTc-HYNIC-DAPI.
HPLC trace of the radioactivity 60(3.6%). The signals from 8.49 to 12.88 min represent different 99mTc-HYNIC-DAPI derivatives (96.4%). After 12 h, the peak at 1.33 min increased to 100%.
Figure 3.
Dose-Response of 99mTc-pertechnetate and 99mTc-HYNIC-DAPI.
Dose-dependent decrease in supercoiled DNA exposed to 99mTc-pertechnetate or 99mTc-HYNIC-DAPI for 2 (A) or 24 hours (B). The dose-dependent increase in open circular DNA was similar for both radiotracers, but the fraction of linear DNA was greater in the presence of 99mTc-HYNIC-DAPI compared with 99mTc-pertechnetate.
Table 2.
Mean numbers of DSBs and SSBs induced by 99mTc pertechnetate and 99mTc-HYNIC-DAPI in the absence or presence of DMSO.
Figure 4.
Example images of agarose gel indicating the effect of DMSO.
Representative images of agarose gels in the absence (A) or presence (B) of DMSO. The plasmid DNA samples were irradiated for 24 h with 50–1250 MBq/mL 99mTc-HYNIC-DAPI. The influence of DMSO on DNA damage (OC: open circular, L: linear, SC: super coiled plasmid DNA fraction) caused by 99mTc-HYNIC-DAPI was compared with that elicited by 99mTc-pertechnetate (99mTcO4−). Notably, the formation of open circular and linear DNA in response to 99mTc-pertechnetate could be prevented by DMSO. The first lane is a non-irradiated plasmid sample, L is a non-irradiated linear plasmid, and M is the marker. The DNA bands at the bottom represent supercoiled DNA.
Figure 5.
Effect of DMSO on plasmid damage.
The influence of the radical scavenger DMSO on the DNA conformational changes caused by 99mTc-pertechnetate and 99mTc-HYNIC-DAPI after 2 (A) or 24 h (B) of irradiation. The formation of open circle and linear DNA in response to 99mTc-pertechnetate was nearly completely suppressed in the presence of DMSO. These data indicated that the plasmid damage by 99mTc-pertechnetate was primarily caused by radicals. There was no noticeable effect of DMSO in the 99mTc-HYNIC-DAPI-treated samples, indicating that the plasmid damage was caused by direct interactions between the radioactive compound and the DNA.
Figure 6.
Number of DSBs and SSBs per DNA molecule.
Analysis of the mean number of DSBs (A) and SSBs (B) per plasmid molecule as a function of 99mTc-HYNIC-DAPI decays during a 2 h irradiation. The numbers of DSBs and SSBs were calculated based on the fluorescence intensity of the linear and supercoiled DNA fractions in the presence (□) or absence (▪) of DMSO.
Figure 7.
Measurement of the Plasmid DNA-bound radioactivity.
The measurements were performed using a gamma counter, and the activity was decay-corrected based on the application time. The data were obtained from a single experiment. A good linear correlation was observed between the applied activity and the bound activity.
Figure 8.
Fluorescence intensity with respect to DNA-bound activity.
Dependence of the relative fluorescent intensity of open circle and linear plasmid DNA on the DNA-bound activity. A good correlation was observed between the activity and the radiation damage. The higher fluorescent intensity after the 24 h incubation time was due to the increased number of radioactive decays.