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Multiple osteochondromatosis (MO), or EXT1/EXT2-CDG, is an autosomal dominant O-linked glycosylation disorder characterized by the formation of multiple cartilage-capped tumors (osteochondromas). In contrast, solitary osteochondroma (SO) is a non-hereditary condition. EXT1 and EXT2, are tumor suppressor genes that encode glycosyltransferases involved in heparan sulfate elongation. We present the clinical and molecular analysis of 33 unrelated Latin American patients (27 MO and 6 SO). Sixty-three percent of all MO cases presented severe phenotype and two malignant transformations to chondrosarcoma (7%). We found the mutant allele in 78% of MO patients. Ten mutations were novel. The disease-causing mutations remained unknown in 22% of the MO patients and in all SO patients. No second mutational hit was detected in the DNA of the secondary chondrosarcoma from a patient who carried a nonsense EXT1 mutation. Neither EXT1 nor EXT2 protein could be detected in this sample. This is the first Latin American research program on EXT1/EXT2-CDG.

Homocystinuria due to CBS deficiency is a rare autosomal recessive disorder characterized by elevated plasma levels of homocysteine (Hcy) and methionine (Met). Here we present the analysis of 22 unrelated patients of different geographical origins, mainly Spanish and Argentinian. Twenty-two different mutations were found, 10 of which were novel. Five new mutations were missense and five were deletions of different sizes, including a 794-bp deletion (c.532-37_736 + 438del794) detected by Southern blot analysis. To assess the pathogenicity of these mutations, seven were expressed heterologously in Escherichia coli and their enzyme activities were assayed in vitro, in the absence and presence of the CBS activators PLP and SAM. The presence of the mutant proteins was confirmed by Western blotting. Mutations p.M173del, p.I278S, p.D281N, and p.D321V showed null activity in all conditions tested, whereas mutations p.49L, p.P200L and p.A446S retained different degrees of activity and response to stimulation. Finally, a minigene strategy allowed us to demonstrate the pathogenicity of an 8-bp intronic deletion, which led to the skipping of exon 6. In general, frameshifting deletions correlated with a more severe phenotype, consistent with the concept that missense mutations may recover enzymatic activity under certain conditions.