Efficiency of siRNA in silencing Braf-V600E mutated osteoclasts in a mouse model of histiocytosis-like disease with bone involvement
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Abstract
Histiocytosis is a group of disorders characterized by the accumulation of aberrant hemopoietic cells that result in inflammatory lesions capable of subsequent damage in multiple organ systems such as the liver, lungs, lymph nodes, spleen, and bone. This disorder results from a mutation in the BRAF gene, a serine/threonine protein kinase that is involved in intracellular signaling pathways critical for cell proliferation and differentiation as well as survival. These mutations result in constitutive activation of the BRAF protein, causing aberrant hyperactivation of this signaling pathway. Langerhans Cell Histiocytosis is a disease resulting from aberrant Langerhans cells affecting multiple organs, including skin, lung, brain, bone, and so forth. Patients that have LCH with bone involvement present with an accumulation of hyperactive osteoclasts and bone resorbing cells, resulting in painful osteolytic lesions across the skeletal system. Current treatments such as BRAF and osteoclast inhibitors demonstrate the capability to manage the disease in the short term but not without long-term side effects, such as drug toxicity or growth defects on longitudinal bone, that predominantly affect children. Our laboratory is working on testing gene silencing (siRNA) of the BRAF mutation to restore normal bone resorption in mouse models of LCH. Our preliminary results first focused on establishing optimal concentrations of siRNA (control) in vitro in osteoclast cultures from wild-type or Braf-mutated bone marrow osteoclast progenitor cells. Then we tested siRNA against the Braf-V600E mutation in these cultures. Results demonstrated that, unlike the control, siRNA-BrafV600E cytotoxicity is higher when compared to the same concentrations utilized for the siRNA control; therefore, siRNA-BrafV600E treatment warrants future investigation.
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2025