Structure-aided design of inhibitors of Mycobacterium tuberculosis thimydilate kinase

SUMMARY The aim of this work was the search for selective inhibitors of M. tuberculosis thymidine monophosphate kinase (TMPKmt) as leads for the development of new anti-tuberculosis agents. Based on the X-ray structure of TMPKmt, it was decided to establish a preliminary structure-activity relationship by synthesising a series of nucleosides and nucleotides modified at the 2’-, 3’- and 5-positions of the dTMP-scaffold. These analogues were tested for their affinities for TMPKmt. From this, a 2'-chlorine and a 2'-fluorine substituent emerged as the most promising modifications of the dTMP scaffold. Furthermore, the affinities of a series nucleosides and their corresponding nucleotides were compared. The deletion of the phosphate moiety typically resulted in a modest, in many cases negligible affinity loss. In view of the drug delivery problems of phosphorylated compounds, nucleosides seemed more useful leads for further drug design. The goal of the synthesis of a series 3'-C-branched-chain nucleosides and nucleotides (3'-CH2N3, 3'-CH2NH2, 3'-CH2F, 3'-CH2OH) was to occupy a cavity in the enzyme near the 3'-position. Biological results and modeling confirmed this hypothesis. 3'-Azidomethyl- 3'-deoxy-thymidine (4.9), combining a low Ki-value (40 mM) with a high selectivity index for the tuberculosis enzyme (Ki TMPKh/ Ki TMPKmt = 26) emerged as the most promising lead for further optimisation. Attempts were made to combine these favourable 3'-substitution patterns with 2'-halogen substituents (6.9-6.14). However, introduction of the 2'-halogen led to a drastic decrease in affinity compared to the corresponding 2'-deoxy-nucleosides. Probably, the 2'-halogens compete with the 3'-substituents for the same binding pocket. Furthermore, it was tried to supersede the good affinities of the 3'-C-branched-chain nucleosides, by further exploration of the enzyme cavity near the 3'-position with alternative nitrogen-containing substituents. However, attempted simultaneous reduction of the 6'-azido function and the 2'-hydroxyl of compound 19 (chapter 6) failed. Instead three peculiar nucleoside analogues (6.10, 6.11 and 6.12) were isolated that were tested for their affinities for TMPKmt and TMPKh. Dinucleoside 6.12 unexpectedly showed a high affinity (Ki = 37 mM) and selectivity for the tuberculosis enzyme, indicating an exceptional flexibility of TMPKmt towards the orientation of the sugar ring. Also the bicyclic nucleosides 6.10 and 6.11 showed excellent affinities. With its Ki-value of 3.5 mM (exceeding the affinity of the natural substrate) and its selectivity index of 200, 6.10 represents the highest affinity and most selective inhibitor of TMPKmt found so far. The favourable affinities and selectivities of these three inhibitors will form the basis for further drug design, considerably increasing the variety of nucleoside analogues that may be envisaged for future synthesis. Based on the encouraging affinity of anhydro-hexitol nucleotide 7.1, the effects of sixmembered sugar rings for the affinity for TMPKmt were further explored through the synthesis of 1-[2,4-dideoxy-4-C-hydroxymethyl-a-L-lyxopyranosyl]thymine (7.4). This nucleoside indeed showed the predicted equatorial orientation of the thymine ring (4C1). Affinity results were however disappointing. In conclusion, this work yielded some high affinity and selective inhibitors for TMPKmt that open interesting perspectives in the search for new anti-tuberculosis agents. Based on their high selectivity indices, especially dinucleoside 6.12 and bicyclic nucleosides 6.10 and 6.11 will be used as the starting point for the search of inhibitors with an optimal fitting in the active pocket of TMPKmt.

Author dc:creator

• Vanheusden, V

Contributors dc:contributor

• Van Calenbergh, S

Statement dc:rights

• info:eu-repo/semantics/openAccess

Language dc:language

und

Identifier

https://biblio.ugent.be/publication/470974
http://doi.org/1854/3511
https://biblio.ugent.be/publication/470974/file/1875793

OAI identifier oai:identifier

oai:archive.ugent.be:470974