Abstract
Accumulating evidence suggests an unequivocal role of lysyl oxidases as key players of tumor progression and metastasis, which renders this enzyme family highly attractive for targeted non-invasive functional imaging of tumors. Considering their function in matrix remodeling, malignant melanoma appears as particularly interesting neoplasia in this respect. For the development of radiotracers that enable PET imaging of the melanoma-associated lysyl oxidase activity, substrates derived from the type I collagen α1 N-telopeptide were labeled with fluorine-18 using N-succinimidyl 4-[18F]fluorobenzoate ([18F]SFB) as prosthetic reagent. With regards to potential crosslinking to tumor-associated collagen in vivo, their interaction with triple-helical type I collagen was studied by SPR. A mouse model of human melanoma was established on the basis of the A375 cell line, for which the expression of the oncologically relevant lysyl oxidase isoforms LOX and LOXL2 was demonstrated in Western blot and immunohistochemical experiments. The radiopharmacological profiles of the peptidic radiotracers were evaluated in normal rats and A375 melanoma-bearing mice by ex vivo metabolite analysis, whole-body biodistribution studies and dynamic PET imaging. Out of three 18F-labeled telopeptide analogs, the one with the most favorable substrate properties has shown favorable tumor uptake and tumor-to-muscle ratio. Lysyl oxidase-mediated tumor uptake was proven by pharmacological inhibition using β-aminopropionitrile and by employing negative-control analogs of impeded or abolished targeting capability. The latter were obtained by substituting the lysine residue by ornithine and norleucine, respectively. Comparing the tumor uptake of the lysine-containing peptide with that of the non-functional analogs indicate the feasibility of lysyl oxidase imaging in melanoma using substrate-based radiotracers.
Highlights
Malignant melanoma is a highly aggressive and treatmentresistant malignancy of melanocytes, which occurs primarily in fair-skinned populations (Tandler et al, 2012)
Radiolabeled lysine-containing peptidic substrates should spontaneously react with lysine residues on extracellular tumor-associated proteins after conversion to allysine by lysyl oxidase
Irrespective of such a scenario, substrate-based radiotracers can be suitable for imaging of enzymes in vivo even in the absence of irreversible product trapping, as demonstrated for positron emission tomography (PET) imaging of histone deacetylases (Seo et al, 2013; Bonomi et al, 2015)
Summary
Malignant melanoma is a highly aggressive and treatmentresistant malignancy of melanocytes, which occurs primarily in fair-skinned populations (Tandler et al, 2012). Proteases, which lead to the partial degradation of proteinaceous ECM components, play a pivotal role in that process (Friedl and Wolf, 2008), and enzymes whose actions result in more subtle structural changes involving the amino acid side chains of proteins are important in that context. Among the latter category of enzymes, lysyl oxidases have been shown to be extraordinarily relevant for the metastasis of solid tumors (Perryman and Erler, 2014)
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