Synthesis of a Small Library of 3-(Carboranylalkyl)thymidines and Their Biological Evaluation as Substrates for Human Thymidine Kinases 1 and 2

Abstract

A small library consisting of two series of thymidine derivatives containing o-carboranylalkyl groups at the N-3 position was prepared. In both series, alkyl spacers of 2-7 methylene units were placed between the o-carborane cage and the thymidine scaffold. In one series, an additional dihydroxypropyl substituent was introduced at the second carbon atom of the carborane cage. In the series of N-3-substituted carboranyl thymidines without additional dihydroxypropyl substituent, three steps were required to obtain the target compounds in overall yields as high as 75%, while in the series of N-3-substituted carboranyl thymidines with additional dihydroxypropyl substituent, 9-10 steps were necessary with significantly lower overall yield. All target compounds were good substrates of human cytosolic thymidine kinase 1 while they were, if at all, poor substrates of the mitochondrial thymidine kinase 2. There was only a minor difference in phosphorylation rates between N-3-substituted carboranyl thymidines with additional dihydroxypropyl substituents with thymidine kinase 1 (range: 13-49% relative to thymidine) and their counterparts lacking this group (range: 11-57% relative to thymidine). Tether lengths of two and five methylene groups in both series gave the highest enzyme activities in the present study. A hypothesis for this result is presented.

Structures of thymidine kinase 1 of human and mycoplasmic origin

“…It has been shown that TK1 accepts large substitutions at the 3-position of the ring (56). Although N3 is hydrogen-bonded to a main-chain carbonyl in this position, the 3-position of the ring points toward the contact area between the ␣͞␤-domain and the lasso domain.…”

Boron-Containing Nucleosides as Potential Delivery Agents for Neutron Capture Therapy of Brain Tumors

“…However, even if the nucleosides only were localized intracellulary rather than incorporated into DNA, they still would be attractive boron delivery agents. Since the mid1990s, more than 40 N-3 substituted thymidine analogs, boronated and non-boronated, have been synthesized by applying concepts originally developed by Soloway et al (5,23) and subsequently pursued by Lunato et al (43) and Tjarks et al (24,44,58). These carboranyl thymidine derivatives have undergone an in-depth enzymatic evalu- Abbreviations: i.c., intracerebral; Tk, thymidine kinase; CED, convection-enhanced delivery; DCP-AES, direct current plasma-atomic emission spectroscopy.…”

“…Fig. 1 summarizes a selection of boron-containing nucleosides, the synthesis of which has been described recently in detail by us (44). These were evaluated in a phosphorylation transfer assay as described previously (44).…”

“…1 summarizes a selection of boron-containing nucleosides, the synthesis of which has been described recently in detail by us (44). These were evaluated in a phosphorylation transfer assay as described previously (44). Briefly, the assay reaction mixture contained 50 mM Tris-HCl (pH 7.6), 5 mM MgCl 2 , 125 mM KCl, 10 mM DTT, 0.5 mg/ml BSA, 1 mM ATP with 0.3 M [␥-…”

“…Therefore, thymidine kinase-1 and 2Ј-deoxycytidine kinase seem to be the primary targets of boronated nucleosides (39,40), and knowledge of their substrate specificity has been indispensable for the rational design of boron-containing nucleoside derivatives (41)(42)(43). Recently, we have synthesized and have carried out a preliminary biochemical evaluation of a small library of boronated thymidine analogs, which revealed that they were excellent substrates of thymidine kinase-1 (44).…”