الفهرس 
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Abstract
Multiple myeloma (MM) is a tumor of plasma cells affecting bone marrow. Elderly are mainly the victims of MM. The disease causes abnormal immunoglobulins production and wide range of complications up to fatal renal failure. Despite the progress in treatments, MM remains intractable disease with poor prognosis.
MM accounts for 10% of all hematological cancers. At the era between 1990 - 2016, incident cases of myeloma have increased by 126% globally.
In Egypt, the incidence rates in 2014 per100, 000 population of MM cancer in lower, middle, and upper Egypt were 0.51%, 0.61% and 0.19%, respectively.
New era in molecular biology has been developed after the discovery of gene editing by clustered regularly interspaced short palindromic repeats (CRISPR) technology. That discovery becomes a hot spot technology used in tremendous biomedical researches.
CRISPR-Cas9 system consists of two key molecules; Cas 9 endonuclease which is the cutter protein and guide RNA which designed to be sequence specific to guide Cas protein into precise nucleotide sequence to produce double strand DNA breaks. The breaks initiate cells repair mechanism to fix it either with knockout or knock in depending on the availability of template DNA, respectively.
This study was performed at Medical Biochemistry and Molecular Biology Department, Faculty of Medicine, Ain Shams University during the period from May 2017- May 2019.
The aim of the present study was to study the effect of gene editing on multiple myeloma cells’ growth. We exploited bioinformatics to retrieve a potential target involved in MM pathogenesis and knocking out that target using CRISPR-Cas9 technology. VpreB1 (CD179a) gene was the selected gene that probably involved in molecular pathogenesis of MM. CRISPR-Cas9 tool was designed to knockout VpreB1 gene from primary myeloma cell line as a therapeutic approach. Assessments were performed to figure out the outcome on viability, count and proliferation of myeloma cells.
We designed gRNA to knockout VpreB1 gene from primary myeloma cell line. The transfection of myeloma cells with gRNA was carried out via two approaches; direct gRNA transfection (approach1) and indirect transfection after cloning of gRNA using plasmid vector (approach2).
Knockout efficiency was assessed using qPCR to detect VpreB1 mRNA relative expression. The VpreB1 protein expression was also assessed by immunofluorescence.
While the effect of VpreB1 gene knockout on myeloma cells was assessed on different parameters. Cell count with trypan blue assay and cell viability and proliferation inhibition with MTS assay were performed on myeloma cells transfected with CRISPR-Cas9 to knockout VpreB1 gene.
The results of our study support our hypothesis that VpreB1 gene is probably involved in MM pathogenesis. That is why after VpreB1 knockout in myeloma cells, the count of viable myeloma cells and percent of viability have markedly decreased, as compared to mock and untreated myeloma control cells.
We also figured out that, approach 1 was better than approach 2 in knockout efficiency. Both VpreB1 gene and protein expressions were lower in myeloma cells transfected via approach 1. Therefore, VpreB1 gene could be a potential therapeutic target for CRISPR-Cas9 gene editing technique to control the proliferation of myeloma cells in patients with multiple myeloma, although extended studies are required.