Anti-SPRED1 monoclonal antibody, clone 573DU3.6.2

A 4-year-old child with no medical history presented for evaluation of a small, palpable nodule near the left inferolateral rim. The lesion had a bluish hue and had been slowly enlarging over the course of several months. MRI of the orbits revealed a heterogenous and infiltrative preseptal and extraconal mass which enhanced with gadolinium, and sphenoid wing dysplasia on the left. A complete ophthalmic and physical examination failed to reveal any other stigmata of neurofibromatosis type 1 (NF1) or neurofibromatosis type 2 (NF2). An incisional biopsy of the palpable mass revealed a plexiform neurofibroma. Molecular sequencing of the NF1 and NF2 genes did not reveal causative mutations. Further investigation revealed a loss of function mutation in SPRED1 on chromosome 15. Although loss of function mutations in the SPRED1 gene are known to cause several dermatologic changes associated with the NF 1-like phenotype, to our knowledge, this is the first description of a SPRED1 gene mutation resulting in ophthalmic abnormalities.

Neurofibromin and Sprouty-related EVH1 domain-containing protein 1 (Spred1) both act as negative regulators of the mitogen-activated protein kinase pathway and are associated with the rare diseases Neurofibromatosis type 1 and Legius syndrome, respectively. Spred1 recruits the major GTPase activating protein (GAP) neurofibromin from the cytosol to the membrane in order to inactivate the small G protein Ras. These functions are dependent on the N-terminal EVH1 domain and the C-terminal Sprouty domain of Spred1 whereas the former specifically recognizes the GAP related domain of neurofibromin and the latter is responsible for membrane targeting. Within the GAP domain, Spred1 binding depends on the GAPex portion which is dispensable for Ras inactivation. In a first step towards the characterization of the Neurofibromin Spred1 interface in solution we assigned backbone and side chain H-1, C-13, and N-15 chemical shifts of the Spred1 derived EVH1 domain. Our chemical shift data analysis indicate seven consecutive beta-strands followed by a C-terminal alpha-helix which is in agreement with the previously reported crystal structure of Spred1(EVH1). Our data provide a framework for further analysis of the function of patient-derived mutations associated with rare diseases.