Moreover, systemic administration of such macromolecules can elicit side reactions in different patients, a risk that may limit their use in humans. To overcome the above limitations, Urano et al.16 recently reported a new class of activatable molecular probes that combine rapid fluorescence enhancement in tumors with high specificity. Taking advantage of the overexpression of γ-glutamyltranspeptidase (GGT) on the cell surface of Z-VAD-FMK in vitro diverse tumors, the authors developed fluorogenic substrates for GGT. They synthesized a spirocyclic γ-glutamyl hydroxymethyl rhodamine
green (gGlu-HMRG) and explored its use as a fluorogenic substrate for GGT. Upon interaction with GGT, cleavage of the glutamyl group results Neratinib in the spontaneous conversion of the structurally constrained spirocyclic molecules to the highly fluorescent hydroxymethyl rhodamine green (HMRG) derivative (Fig. 1). Because GGT is a cell surface enzyme, and the substrate is a small molecule, the conversion of spirocyclic gGlu-HMRG to HMRG would be expected to result in rapid clearance of the fluorophore from the tumor site, a condition that would lead to nonspecific or low fluorescence in the region
of interest. Instead, the authors observed highly localized fluorescence signal in tumors. This fortuitous observation was attributed to the generation of a more hydrophobic HMRG that rapidly internalizes in cells than the relatively hydrophilic gGlu-HMRG that is less permeable to cell membranes. Although this is this website a reasonable explanation, additional studies are needed to delineate the exact mechanism of HMRG’s cellular uptake. A closer examination of the cellular and in vivo images shows that the fluorescence emanates almost exclusively from the intracellular compartment. These data suggest the possibility of a multistep activation pathway, where the spirocyclic structure remains intact
after rapid cleavage of the γ-glutamate, followed by internalization of the more hydrophobic non-fluorescent spirocyclic HMRG, which subsequently isomerized to the highly fluorescent “open form” HMRG in the acidic lysosomes. Regardless of the mechanism of internalization, the specificity and rapid activation of gGlu-HMRG by tumors represents a major advance in the fields of molecular imaging and image-guided surgery. An important outcome of the rapid and specific fluorescence activation is the potential to develop aerosolized activatable molecular probes for topical application in the surgical field. Urano et al.16 showed that small tumor nodules are identifiable within 10 seconds of spraying the activatable molecular probe gGlu-HMRG. For surgical guidance, the topical application of the molecular probe has several advantages, including the elimination of systemic toxicity and the use of small amounts of the molecular probes.