Ectopic calcification in pseudoxanthoma elasticum responds to inhibition of tissue-nonspecific alkaline phosphatase

Abstract

The ABCs of PXE Pseudoxanthoma elasticum (PXE) is a genetic disorder caused by mutations in ABCC6 that is characterized by calcium deposition outside of the skeletal system, specifically in the blood vessels, skin, and eyes. Using patient-derived fibroblasts and genetic knockout mouse models, Ziegler et al. demonstrate that ABCC6 mutant cells are osteogenic and that loss of ABCC6 reduces pyrophosphate, an inhibitor of calcification. In mice, ectopic calcification was seen only when ABCC6 was deleted jointly from the liver and from Wnt1+ cells, suggesting systemic and local contributions to the phenotype. Treating mice and cells with a tissue-nonspecific alkaline phosphatase (TNAP) inhibitor prevented pyrophosphate degradation and ectopic calcification progression. Biallelic mutations in ABCC6 cause pseudoxanthoma elasticum (PXE), a disease characterized by calcification in the skin, eyes, and blood vessels. The function of ATP-binding cassette C6 (ABCC6) and the pathogenesis of PXE remain unclear. We used mouse models and patient fibroblasts to demonstrate genetic interaction and shared biochemical and cellular mechanisms underlying ectopic calcification in PXE and related disorders caused by defined perturbations in extracellular adenosine 5′-triphosphate catabolism. Under osteogenic culture conditions, ABCC6 mutant cells calcified, suggesting a provoked cell-autonomous defect. Using a conditional Abcc6 knockout mouse model, we excluded the prevailing pathogenic hypothesis that singularly invokes failure of hepatic secretion of an endocrine inhibitor of calcification. Instead, deficiency of Abcc6 in both local and distant cells was necessary to achieve the early onset and penetrant ectopic calcification observed upon constitutive gene targeting. ABCC6 mutant cells additionally had increased expression and activity of tissue-nonspecific alkaline phosphatase (TNAP), an enzyme that degrades pyrophosphate, a major inhibitor of calcification. A selective and orally bioavailable TNAP inhibitor prevented calcification in ABCC6 mutant cells in vitro and attenuated both the development and progression of calcification in Abcc6−/− mice in vivo, without the deleterious effects on bone associated with other proposed treatment strategies.

Inhibition of Tissue-Nonspecific Alkaline Phosphatase Attenuates Ectopic Mineralization in the Abcc6 Mouse Model of PXE but Not in the Enpp1 Mutant Mouse Models of GACI

“…Further studies might require higher doses for more drastic reduction in TNAP activity and complete mineralization arrest. Please note that the investigational drug, the dose at 75 mg/ kg/day, and the Abcc6 e/e mouse model for PXE in this study mirrors those used in another study (Ziegler et al, 2017). Combined results of these independent and complementary studies suggested that inhibition of TNAP by SBI-425 could be turned into a viable approach for treatment of patients with PXE.…”

Generation and Validation of a Complete Knockout Model of abcc6a in Zebrafish

“…Its documented expression in chondrocytes and tibia may, however, suggest ABCC6 to be also relevant in human osteoblasts (Beck et al, 2005;Kolesnikov et al, 2015). As such, our knockout model will also be valuable for elucidation of the complex pathomechanisms underlying PXE, the osteogenic changes in particular, as studies suggest that the aberrant inorganic pyrophosphate release that was identified is not the only mechanism leading to ectopic mineralization in PXE (Zhao et al, 2017;Ziegler et al, 2017).…”

Comparison of inbred mouse strains shows diverse phenotypic outcomes of intervertebral disc aging

“…While studies of single gene knockouts affecting PPi metabolism showed ectopic mineralization of the AF and adjacent ligaments (Warraich et al, 2013; Zhang et al, 2016) LG/J mice recapitulated age‐dependent disc calcification that affected both the NP and AF similar to elderly humans (Chanchairujira et al, 2007). Studies have also shown a causal relationship between systemic activity of TNAP that degrades PPi into Pi and ectopic calcification of soft tissues (Ziegler et al, 2017). Interestingly, blood analysis showed that several mineral metabolism markers including free Ca 2+ and phosphorous were not affected by the strain background.…”