Bioconjugation of holo-transferrin with hypoxia-enhanced fluorescent sensor for the selective imaging of cancer cells

A new fluorescent sensor for imaging cancer cells was designed by bioconjugation of holo-transferrin (hTf) with a nitro-pyrazinotriazapentalene moiety (PyTAP-NO2) as a bioreductive fluorescent unit. hTf was chosen to increase its selective delivery to cancer cells through transferrin receptor (TfR) mediated endocytosis. PyTAP-NO2 was applied to enhance the fluorescent signal in oxygen-deprived cells due to the increased expression of nitroreductases under such conditions. A protein-dye bioconjugate had an average of 8 fluorophore molecules per protein that effectively amplifying the fluorescence signal. The designed hTf- PyTAP-NO2 was non-toxic and its efficient accumulation through TfR, as well as its high stability and sustained intracellular retention, was proven. hTf-PyTAP-NO2 bioconjugate allows selective accumulation in cancer cells containing increased levels of TfRs with simultaneous slight enhancement of signal in hypoxic cells due to increased expression of nitroreductases.