Raffaele is part of a joint collaboration project between Fran Platt’s (Oxford) and Andrea Ballabio’s (TIGEM, Italy) lab studying Niemann-Pick disease type-C 1 and 2.

Niemann-Pick type C (NP-C) is a lysosomal lipid storage disorder caused by autosomal recessive mutations in either the NPC1 or NPC2 genes. The majority of the NPC cases, about 95%, results from dysfunction of NPC1, whereas the remaining 5% is due to malfunction of NPC2. Clinically, NPC is a neurovisceral disorder characterized by a progressive neurodegeneration similar to Alzheimer’s disease.

At the cellular level, upon sphingosine accumulation, Ca²⁺ homeostasis is altered leading to cholesterol, sphingomyelin and glycosphingolipids (GSLs) build-up within NPC cells. To date, the mechanisms linking lipid storage and onset of organ dysfunction are still unknown. In this respect, a major limitation for in vitro studies is represented by the lack of appropriate cellular models of NPC.

Therefore, in the Ballabio lab using CRISPR/Cas9 technology, I knocked-out either the NPC1 or the NPC2 genes in ARPE19 cells, obtaining isogenic cellular models of NPC type 1 and type 2, respectively. While based in the Platt lab, I have characterised NPC1^-/- ARPE19 and NPC2^-/- ARPE19 demonstrating that they exhibit the classical spectrum of the pathological phenotypes of Niemann-Pick C. I am currently using these cell lines to study the basic biology of the two forms of NPC.

Key Points:

• Comparative study of NPC type1 and type2 through the characterization of the newly generated NPC-cellular models, NPC1^-/- ARPE19 and NPC2^-/-
• Investigation of the physiology of calcium in NPC2^-/- ARPE19 to assess whether the levels of lysosomal calcium in these cells are altered with respect to NPC1^-/- ARPE19 and healthy controls.
• In vitro testing of potential therapeutic compounds monitoring their effects on the multiple NPC-like phenotypes shown by the NPC1^-/- and the NPC2^-/- ARPE19 cell lines.