L improve cellular selectivity of deep tissue PDT, thus increasing the probability of complete tumor cell eradication, whilst retaining the viability of healthy tissue. Activatable antibody-based probes merge the concept of targeted PS delivery with tumor-selective activation, which is another mechanism for a class of targeted PDT probes that will be discussed in the next section.Degree of probe conjugationImportantly for PDT, the number of PSs conjugated per targeting probe has also been shown to exert a marked impact on its pharmacokinetics. Vrouenrats et al. investigated the pharmacokinetic behavior of a radiolabeled chimeric Ig anti-epican antibody, 125I-cMAb U36, conjugated to an amine-reactive tetrafluorophenyl ester of the porphyrin derivative TrisMPyPFCO2H [144]. The antibody-to-PS ratio was varied from 1:1.2, 1:2.1 and 1:3.0; the PIC was unstable and aggregated when 4 PS molecules were conjugated to each antibody. Tumor uptake was reduced when more PSs were conjugated to each antibody as HNX-OE head and neck mouse tumors were found to have 15.5, 8.6, 6.5, and 4.0 injected dose (ID)/g tissue at 48 h following administration of the unconjugated, 1:1.2, 1:2.1 and 1:3.0 PICs, respectively. The reduced tumor delivery could be attributed to decreasing circulation half-lives as the degree of PS conjugation increased. At 48 h following injection, the 1:1.2, 1:2.1 and 1:3.0 PICs were found in the blood at 58 , 42 , and 26 of the unconjugated antibody in circulation, respectively. The expedited clearance rates with increasing degree of PS conjugation to the PICs did not correspond to higher tumor-to-normal ratios; on the contrary an inversely proportional relationship was observed. Interestingly, it has been shown that antibody-mediated agent delivery in tumors does not increase the total tumor uptake of the probes, as a sham non-selective IgG antibody accumulates in the tumor to the same degree as a tumor-specific IgG [136]. However, a stark difference in receptor binding and therapeutic efficacy is observed between the targeted antibody and the non-selective IgG. Activatable variants of the BPD-cetuximab PIC were developed by Savellano et al. who demonstrated that an increase in the degree of PS conjugation to the antibodies increased the static quenching of the BPD, reducing the photoactivity [145]. Spring et al., then utilized the quenched 7:1 BPD:cetuximab PICs, which exhibited a 7-fold quenching in photoactivity whilst intact, for the selective, tumor-specific activatable PDT of ovarian micrometastasis [146]. The activatable PIC leverages the specific cellular discrimination of antibody conjugates, regardless of total tumor concentration. The capacity for the tumor-activatable PIC to become activated following specific cancer cell binding, internalization and a proteolytic degradation adds further selectivity to PDT treatments that EPZ004777 cancer utilize targeted probes. In the presence of theProbes for selective tumor activation Photodynamic molecular beaconsSelectivity towards the target tissue that will be treated with PDT can be also imparted through the inherent DM-3189 mechanism of action biochemical lability of customized probes that are designed to be enzymatically or chemically cleaved, and thus activated, at the required site of action (Fig. 6B). Therefore, targeted PDT is no longer confined by selective tumor delivery and localized photo-irradiation, but is also regulated by tumor biochemistry and microenvironment. An archetypal example of such activatable tec.L improve cellular selectivity of deep tissue PDT, thus increasing the probability of complete tumor cell eradication, whilst retaining the viability of healthy tissue. Activatable antibody-based probes merge the concept of targeted PS delivery with tumor-selective activation, which is another mechanism for a class of targeted PDT probes that will be discussed in the next section.Degree of probe conjugationImportantly for PDT, the number of PSs conjugated per targeting probe has also been shown to exert a marked impact on its pharmacokinetics. Vrouenrats et al. investigated the pharmacokinetic behavior of a radiolabeled chimeric Ig anti-epican antibody, 125I-cMAb U36, conjugated to an amine-reactive tetrafluorophenyl ester of the porphyrin derivative TrisMPyPFCO2H [144]. The antibody-to-PS ratio was varied from 1:1.2, 1:2.1 and 1:3.0; the PIC was unstable and aggregated when 4 PS molecules were conjugated to each antibody. Tumor uptake was reduced when more PSs were conjugated to each antibody as HNX-OE head and neck mouse tumors were found to have 15.5, 8.6, 6.5, and 4.0 injected dose (ID)/g tissue at 48 h following administration of the unconjugated, 1:1.2, 1:2.1 and 1:3.0 PICs, respectively. The reduced tumor delivery could be attributed to decreasing circulation half-lives as the degree of PS conjugation increased. At 48 h following injection, the 1:1.2, 1:2.1 and 1:3.0 PICs were found in the blood at 58 , 42 , and 26 of the unconjugated antibody in circulation, respectively. The expedited clearance rates with increasing degree of PS conjugation to the PICs did not correspond to higher tumor-to-normal ratios; on the contrary an inversely proportional relationship was observed. Interestingly, it has been shown that antibody-mediated agent delivery in tumors does not increase the total tumor uptake of the probes, as a sham non-selective IgG antibody accumulates in the tumor to the same degree as a tumor-specific IgG [136]. However, a stark difference in receptor binding and therapeutic efficacy is observed between the targeted antibody and the non-selective IgG. Activatable variants of the BPD-cetuximab PIC were developed by Savellano et al. who demonstrated that an increase in the degree of PS conjugation to the antibodies increased the static quenching of the BPD, reducing the photoactivity [145]. Spring et al., then utilized the quenched 7:1 BPD:cetuximab PICs, which exhibited a 7-fold quenching in photoactivity whilst intact, for the selective, tumor-specific activatable PDT of ovarian micrometastasis [146]. The activatable PIC leverages the specific cellular discrimination of antibody conjugates, regardless of total tumor concentration. The capacity for the tumor-activatable PIC to become activated following specific cancer cell binding, internalization and a proteolytic degradation adds further selectivity to PDT treatments that utilize targeted probes. In the presence of theProbes for selective tumor activation Photodynamic molecular beaconsSelectivity towards the target tissue that will be treated with PDT can be also imparted through the inherent biochemical lability of customized probes that are designed to be enzymatically or chemically cleaved, and thus activated, at the required site of action (Fig. 6B). Therefore, targeted PDT is no longer confined by selective tumor delivery and localized photo-irradiation, but is also regulated by tumor biochemistry and microenvironment. An archetypal example of such activatable tec.