CLL is a malignant disease of B-cells characterised by a heterogeneous clinical outcome. Some patients require an early treatment and have low survival time, while others never need treatment. Many prognostic markers have been established and used to help predict the clinical course of CLL. Despite advances in understanding the biology of CLL, the molecular differences underlying the variable clinical outcome of CLL are not yet fully understood. The hypothesis of this study was that the heterogeneous outcome of CLL could be driven, in part, by the aberrant expression of proteins in the two forms of CLL. Therefore, this study aimed to identify these proteins using proteomics approaches. In an attempt to achieve this goal, four steps were performed. Firstly, a cellular fractionation method was developed to extract cellular proteins into two different fractions (NP40 fraction for cytosolic protein enrichment and SDS fraction for nuclear protein enrichment). Secondly, extracted proteins were subjected to qualitative proteomics analysis using 2D nano-LC and MALDI TOF-TOF mass spectrometry in order to identify CLL proteins. Integrating the identified proteins (n=900) with previously published transcriptome of CLL cells and normal B-cells highlighted 20 proteins with preferential expression in CLL cells - some of which were linked to human cancer. Thirdly, iTRAQ technology coupled with 2D nano-LC and MALDI TOFTOF mass spectrometry was used to measure the relative expression of proteins in different CLL samples. This workflow identified 15 altered proteins in the two forms of CLL and detected 14 proteins with variable expression. Finally, six proteins were selected for investigation in an additional CLL cohort. Of these proteins thyroid hormone receptor-associated protein 3 (TR150), T-cell leukaemia/lymphoma protein 1A (TCL-1) and S100A8 showed association with poor prognosis CLL and early requirement for treatment. Additionally, myosin-9 exhibited reduced expression in poor prognosis CLL samples.! ! Overall, this study identified proteins with potential importance in CLL prognosis and pathology. These proteins merit investigation in a larger CLL cohort to further confirm their relevance to CLL. In addition, this study showed the usefulness of combining cellular fractionation with proteomics and transcriptomics to identify proteins with potential role in CLL.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:571757 |
| Date | January 2013 |
| Creators | Alsagaby, Suliman Abdallah A. |
| Publisher | Cardiff University |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://orca.cf.ac.uk/47568/ |
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