Preparation of 4-alkoxy Substituted Pyrrolopyrimidines for CSF-1R Structure-Activity Relationship Studies

Abstract

Overexpression of the colony stimulating factor 1 (CSF-1R), a tyrosine kinase, is linked to numerous diseases, which can be treated by small molecule inhibitors.[1] Over the course of this master thesis, four new alkoxylated pyrrolopyrimdines, with an ether group at C-4, were synthesized. The final compounds were tested for their activity towards the inhibition of (CSF-1R) and epidermal growth factor receptor (EGFR). First, the pyrrole group of the commercially available 4-chloro-7Hpyrrolo[2,3-d]pyrimidine (3) was protected using 2-(trimethylsilyl)-ethoxymethyl (SEM-Cl, 4) in 24-81% yield. Through directed lithiation C-6 was iodinated to give 4-chloro-6-iodo 7-((2-(trimethylsilyl)ethoxy)methyl)-7H-pyrrolo[2,3d]pyrimidine (7, Scheme 1) in 92-93% yield. Alkoxylations at C-4 were performed on compounds 7 and 13 (13-85%, Scheme 2). Reaction times were reduced by using higher temperatures, but higher temperatures caused decomposition. Alkoxylation using phenol (9) was also investigated in a small study via microwave reactor, but when the same conditions were applied for the reaction with (rac)-2,2,2-trifluoro-1-phenylethan-1-ol ((rac)-19), no product was obtained because of decomposition. Overall, alkoxylations using the trifluorophenylethanols lead to lower yields also due to decomposition. Following the alkoxylation (Route A) Suzuki couplings, to introduce a sp2-sp2 bond at C-6 (yields: 65-73%), and deprotection were used to reach the final compounds (yields: 65-83%). Further work showed that the alkoxylation and Suzuki reactions can be interchanged (Route B). In this pathway, the Suzuki reaction gave 80-81% yield but the alkoxylation reactions initially performed worse achieving 13-70% yield. Overall, yield improvement required a lot of individual fine tuning. In the Suzuki reaction, temperatures of 60 °C gave higher yields, than reactions performed at 100 °C. Both catalyst systems, PdCl2dppf and XPhos 2nd generation, have been applied successfully. The two step deprotection in DCM/TFA and THF/NaHCO3 gave yields between 65-83%. Loss in yields were mostly due to issues during the purification process. For the final compound 23 IC50 values of 1.48 nM for the inhibition of CSF-1R were measured and the inhibition concentration profile is superior to the internal standard PLX3397 (IC50 = 10.57 nM). EGFR is also inhibited by 94% at the maximum test concentration of 500 nM. The enantioenriched compounds (R)-24, (S)-24 showed a low inhibitory effect on both and expressed IC50 values >1000 nM.