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.