Ph.D Thesis

Ph.D StudentChefetz Ilana
SubjectPathomechanisms of Hyperphosphatemic Familial Tumoral
DepartmentDepartment of Medicine
Supervisor PROF. Eli Sprecher
Full Thesis textFull thesis text - English Version


Familial Tumoral Calcinosis (FTC) refers to a group of disorders inherited in an autosomal recessive fashion. Two major FTC types have been described: hyperphosphatemic FTC (HFTC; MIM211900),   which is characterized by hyperphosphatemia leading to the development of large periarticular calcified masses; and normophosphatemic FTC (NFTC; MIM610455), featuring smaller calcified masses located at pressure points. HFTC results from mutations in GALNT3, encoding a glycosyltransferase termed ppGalNac -T3, which is responsible for initiating O-glycosylation, and NFTC results from mutations in SAMD9.

In the present study, we (1) identified novel mutations in HFTC and NFTC patients; (2) investigated the pathogenesis of HFTC and NFTC, revealing biochemical pathways suggesting novel therapeutic strategies.

Soon after the discovery of mutations in GALNT3, it appeared that many individuals, displaying typical features of HFTC, do not carry mutations in this gene. We therefore assessed other candidate genes of potential relevance to phosphate homeostasis. The best studied among these is FGF-23, encoding the fibroblast growth factor 23 (FGF-23), which is a member of the fibroblast growth factors family and functions as a potent phosphaturic protein. A mutation analysis of a Turkish HFTC affected family revealed a homozygous T>C transversion at position 287 of the FGF-23 cDNA sequence. It is now known that FGF-23 undergoes ppGalNac -T3-mediated O-glycosylation, which is necessary to protect it from proteolysis, providing a mechanistic basis for our genetic findings.

We assessed the effects of GALNT3 down-regulation on dermal fibroblasts in an in vitro setting in isolation from the systemic effects of ppGalNac -T3 and demonstrated that GALNT3 expression is under the regulation of inorganic phosphate, calcium and 1,25-dihydroxyvitamin D3. In addition, we showed that decreased GALNT3 expression leads to increased expression of FGF-7 and of MMPs, which have been previously implicated in the pathogenesis of ectopic calcification.

All patients assessed with NFTC had been previously shown to be of Jewish Yemenite and to carry a single mutation, p.K1495E, in SAMD9. Here, we evaluated another Jewish-Yemenite NFTC kindred and found that all patients were compound heterozygous for two mutations in SAMD9: K1495E and a previously unreported nonsense mutation, R344X, predicted to result in a markedly truncated molecule. Our observations indicate that SAMD9 may be a downstream target of TNF-a signaling through p38 pathway; mediating pro-apoptotic signals or activating counter-regulatory anti-inflammatory activities. Interestingly, sorbitol, which, like TNF-a, signals through p38, induced SAMD9 expression, suggesting a common regulatory pathway for SAMD9, involving inducers of cellular stress and inflammatory cell response.