Synthesis, flow cytometric evaluation, and identification of highly potent dipyridamole analogues as equilibrative nucleoside transporter 1 inhibitors

Abstract

Dipyridamole (Persantine) is a clinically used vasodilator with equilibrative nucleoside transporters 1 and 2 (ENT1 and ENT2) inhibitory activity albeit less potent than the prototype ENT1 inhibitor nitrobenzylmercaptopurine riboside (NBMPR). Dipyridamole is a good candidate for further exploration because it is a non-nucleoside and has a proven record of safe use in humans. A series of dipyridamole analogues were synthesized with systematic modification and evaluated as ENT1 inhibitors by flow cytometry. Compounds with much higher potency were identified, the best being 2,6-bis(diethanolamino)-4,8-diheptamethyleneiminopyrimido[5,4-d]pyrimidine (13) with a K(i) of 0.49 nM compared to a K(i) of 308 nM for dipyridamole. Compound 13 is similar in potency to the prototype potent ENT1 inhibitor NBMPR (0.43 nM). For the first time, a dipyridamole analogue has been identified that is equipotent with NBMPR. The SAR indicated that diethanolamine substituted analogues were more active than monoethanolamine compounds. Also, free hydroxyl groups are not essential for activity.

Figure 1

Representatives of the three main ENT1 inhibitory chemical classes

Figure 2

Structure of SAENTA-fluorescein

Figure 3

Representative regions for dipyridamole analogs

Scheme 1^a

^aReagents and conditiond: (a) NHR¹R², Anhydrous THF, 0 - 5 °C; (b) NHR³R⁴, DMSO, 150 °C; (c) R¹MgCl, Anhydrous THF, 0 - 5 °C.

Scheme 2^a

^aReagents and conditions: (a) HCOOH, 100 °C (compound 74); (b) CH₃COCl, DMAP, anhydrous THF, 0-5 °C (compound 75); (c) NaH, R¹I, anhydrous DMF (R² = R¹ for 76-78; R² = H for 79).