CONTEXT

Mast cells (MCs) are tissue resident cells playing a pivotal role in IgE-dependent allergic reactions or in various inflammatory reactions. MCs are also involved in rare malignancies, mastocytosis, where there are frequently acquired abnormality in the structure of KIT receptor, which binds stem cell factor (SCF), the major cytokine involved in differentiation, proliferation, survival and activation of MCs. Both conditions (allergy and mastocytosis) are currently poorly treated and there is a need to develop cellular models relevant for high-throughput screening of molecules that specifically target either signaling of the high affinity receptor for IgE for allergy, or KIT receptor or its downstream signaling in the context of mastocytosis. However, today, the cell models used in these indications have many limitations: they are not reproducible and costly to produce.
To overcome these limitations, we have isolated a new SCF-dependent human MC line with a fully functional receptor for IgE. In addition, we derived from this KIT Wild-Type (WT) cell line, two subclones expressing the mutated KIT D816V or the mutated KIT Δ417-419 insY, two KIT abnormalities frequently encountered in the course of mastocytosis. 

TECHNICAL DESCRIPTION AND DEVELOPMENT STAGE

The present invention relates to the development of:

• A new SCF-dependent human MC line called ROSA KIT WT with all the morphologic and phenotypic features of normal human MCs. ROSA KIT WT cells can be produced in large amounts and express significant levels of the high affinity receptor for IgE (FceRI). They are activable by aggregation of FceRI, resulting in immediate degranulation and delayed synthesis of inflammatory mediators.
• Two new SCF-independent human MC lines termed ROSA KIT D816V and ROSA KIT Delta 417-419 insY, which can be grown in a medium without SCF, and where mutant KIT is constitutively activated. These two MC lines can be produced in large amounts at low cost and represent a paradigm of the malignant MCs found in mastocytosis patients. They also engraft immuno-deficient NSG mice, reproducing an in vivo model of mastocytosis.

BENEFITS

The benefits are the provision of new human MC lines reflecting at the best the in vivo counterparts, easy to produce in large amounts in a non expensive manner and suitable to to develop high-throughput screening testing in the search of:
1) anti-allergic and / or anti-inflammatory therapeutics.
2) specific inhibitors of different mutations in KIT or inhibitors of signaling pathways initiated by the abnormal mutated KIT receptor

INDUSTRIAL APPLICATIONS

The main industrial applications of this invention are:
1) Large scale studies on the molecular events following FceRI aggregation in human mast cells, leading to identification of new intracellular targets involved in allergy.
2) High-throughput screening of chemical libraries of potential anti-allergic compounds in vitro.
Identification of new targets related to mastocytosis and high-throughput screening of chemical libraries of compounds potentially active on these targets or on mutant KIT in vitro. Compounds selected by this way could thus be tested in vivo in grafted NSG mice.

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