Mechanism of action of SNF472, a novel calcification inhibitor to treat vascular calcification and calciphylaxis

Abstract

Background and Purpose: No therapy is approved for vascular calcification or calcific uraemic arteriolopathy (calciphylaxis), which increases mortality and morbidity in patients undergoing dialysis. Deposition of hydroxyapatite (HAP) crystals in arterial walls is the common pathophysiologic mechanism. The mechanism of action of SNF472 to reduce HAP deposition in arterial walls was investigated. Experimental Approach: We examined SNF472 binding features (affinity, release kinetics and antagonism type) for HAP crystals in vitro, inhibition of calcification in excised vascular smooth muscle cells from rats and bone parameters in osteoblasts from dogs and rats. Key Results: SNF472 bound to HAP with affinity (KD) of 1–10 μM and saturated HAP at 7.6 μM. SNF472 binding was fast (80% within 5 min) and insurmountable. SNF472 inhibited HAP crystal formation from 3.8 μM, with complete inhibition at 30.4 μM. SNF472 chelated free calcium with an EC50 of 539 μM. Chelation of free calcium was imperceptible for SNF472 1–10 μM in physiological calcium concentrations. The lowest concentration tested in vascular smooth muscle cells, 1 μM inhibited calcification by 67%. SNF472 showed no deleterious effects on bone mineralization in dogs or in rat osteoblasts. Conclusion and Implications: These experiments show that SNF472 binds to HAP and inhibits further HAP crystallization. The EC50 for chelation of free calcium is 50-fold greater than a maximally effective SNF472 dose, supporting the selectivity of SNF472 for HAP. These findings indicate that SNF472 may have a future role in the treatment of vascular calcification and calcific uraemic arteriolopathy in patients undergoing dialysis.