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Ligand-Directed Acid-Sensitive Amidophosphate 5‑Trifluoromethyl-2′-Deoxyuridine Conjugate as a Potential Theranostic Agent

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journal contribution
posted on 15.05.2013, 00:00 authored by Tatyana S. Godovikova, Vladimir A. Lisitskiy, Natalya M. Antonova, Tatyana V. Popova, Olga D. Zakharova, Alexey S. Chubarov, Igor V. Koptyug, Renad Z. Sagdeev, Robert Kaptein, Andrey E. Akulov, Vassily I. Kaledin, Valeriy P. Nikolin, Sergei I. Baiborodin, Ludmila S. Koroleva, Vladimir N. Silnikov
Herein, we report a novel strategy to engineer an acid-sensitive anticancer theranostic agent using a vector–drug ensemble. The ensemble was synthesized by directly conjugating the linoleic acid (LA)-modified branched polyethyleneimine with a chemotherapeutic drug trifluorothymidine. Linoleic acid residues were grafted onto 25 kDa polyethyleneimine (PEI) by treating PEI with linoleic acid chloroanhydride. 5-Trifluoromethyl-2′-deoxyuridine (trifluorothymidine, TFT) was introduced into LA-PEI conjugate by phosphorylating the conjugate with amidophosphate of trifluorothymidine 5′-monophosphate (pTFT), which had been activated by its conversion into the N,N-dimethylaminopyridine derivative. The extent of mononucleotide analog incorporation in the polymer was regulated by the ratio of pTFT to the polymer during the synthesis. Samples containing 20–70 TFT residues per PEI molecule were obtained. The cytotoxicity of PEI-LA-pTFT conjugates decreased with increasing nucleotide content, as examined using the MTT method. Due to the presence of fluorine atoms, TFT-based conjugates could be detected directly in the animals by 19F magnetic resonance imaging. In addition, the presence of the amidophosphate group in PEI-LA-pTFT conjugates allowed their detection by in vivo 31P NMR spectroscopy. Indeed, the 31P NMR signal of a phosphoramide (δ ∼ 12 ppm) was observed in the mouse muscle tissue treated with PEI-LA-pTFT conjugate along with the signals from endogenous phosphorus-containing compounds. At the same time, the use of PEI-LA-pTFT conjugate for chemotherapeutic drug delivery is limited due to the low release of pTFT from the carrier. To enhance the release of the drug from the conjugate in the endosomes, PEI-LA polymer was coupled with urocanic acid (UA), which bears imidazole ring and thus can form an acid-labile P–N bond with pTFT. The PEI-LA-UA-pTFT conjugate containing 30 residues of UA and 40 residues of pTFT was tested against the murine Krebs-II ascites carcinoma, grown as an ascetic tumor. The intraperitoneal injection of the conjugates resulted in prolongation of the animals’ life and to the complete disappearance of the tumor after three injections.