Applying RNA Interference Technology to Understand the Role of 1q Amplification in Multiple Myeloma Pathogenesis
Project Summary
Multiple myeloma is the second most common haematological cancer. It is a plasma cell tumour that is characterised by a remarkable molecular heterogeneity and remains an incurable malignancy despite the recent expansion of available treatment options. Primary genetic events create the culprit immortalised plasma cells while the accumulation of secondary genetic events and microenvironment changes derive the malignant transformation. There has been a significant increase in the available therapies for multiple myeloma. However, despite their effectiveness, many are associated with limiting side effects and none of those therapies targets the very genetic events that lead to the malignant transformation of the plasma cells.
Several molecular aberrations collaborate to bring about the malignant transformation of plasma cells. Primary genetic events include trisomies of odd-numbered chromosomes and/or chromosomal translocations involving the immunoglobulin heavy chain. Those changes are not enough, however, to derive the full malignant transformation and hence transformation to multiple myeloma is characterised by the acquisition of secondary genetic events such as chromosome 1 abnormalities which is the main focus of this project. 1q gain is a clonal secondary copy number change that is observed in around 40% of myelomas. 1q gain (3 copies) and amplification (≥4 copies) are associated with increased cytogenetic risk and poorer survival. 1q gain and amplification were associated with a significant reduction in progression-free survival (53 vs 21.8 months respectively). Furthermore, when 1q gain is combined with other high-risk cytogenetic changes was associated with significantly inferior survival.
During the summer placement project, the student will be trained on several state-of-the-art cell culture techniques and cell viability assays that will be tested on a collection of multiple myeloma cell lines with different levels of chromosome 1q gain such as U266 (3 copies), KMS12BM (3-4 copies), MOLP8 (6-8 copies). Using RNA interference technology the student will test the effect of several candidate genes on chromosome 1q that are implicated in the multiple myeloma progression. The silencing effect of those genes will be studied in isolation as well as combined to decipher any synergistic effect. Silencing will be confirmed by quantification at the RNA level using qRT-PCR post-transfection as well as at the protein level using FACS.
The next steps in the project will be in vitro assays to determine the effect of silencing the candidate genes on multiple myeloma cell lines’ growth using various cell viability assays such as Alamar Blue and Apoptotic kits.
Training: Training will be provided in cell culture, RNA extraction, histology, PCR, FACS and data and image analysis.