Defining the cellular origin of chronic lymphocytic leukaemia

Leukaemia is a cancer that results from a disruption in the normal development of cells within the blood. This disruption can occur when regulatory elements responsible for controlling white blood cell development malfunction. We recently discovered that a disturbance in the function of a ubiquitously expressed element protein kinase C during white blood cell development, initiates a specific type of leukaemia, chronic lymphocytic leukaemia (CLL). CLL is the most common adult blood cancer in the Western world, for which there is currently no cure. While many research groups are engaged in investigating the mechanisms that can induce CLL cell death in patients as a mechanism to remove the leukaemic cells, little is known about the origin of CLL cells. Until now it has been almost impossible to address this question, as the factors responsible for this initiating this leukaemia have remained elusive. However, our novel mouse model provides us with a unique opportunity to gain fundamental knowledge about the cellular origin of CLL, and may assist in the generation of novel treatments to cure CLL. Within our university-based research laboratory, we will perform established cellular and molecular techniques to identify which cell is responsible initiating CLL. In parallel with these experiments, we will characterise the CLL-initiating cells in CLL-patient cell samples. In this way, we will uncover important information regarding the cell of origin for CLL.

B-cell chronic lymphocytic leukaemia (CLL) is the most prevalent leukaemia in the Western world with an incidence of 3 per 100,000, for which there is currently no cure. CLL cells morphologically resemble mature small B lymphocytes with a characteristic immunophenotype (CD5+CD19hiCD23+mIgMlo), a resistance to apoptosis, with up-regulation of anti-apoptotic proteins such as Bcl-2 and Mcl-1, and cell cycle arrest at the G0/G1 phase of the cell cycle. Clinically, CLL is a heterogeneous disorder in which some patients survive over a decade with stable disease, while in others the leukaemia behaves aggressively with survival measured in months. Over the last decade, several markers of poor prognostic significance have been identified, however to date no single genetic event has been associated with the initiation of CLL. In addition, the cell of origin of CLL has not been formally identified, although it is thought to originate from a maure B cell population, due to phenotypic and gene expression profiles. In an effort to address this question, we developed a unique mouse CLL model. This was achieved by retrovirally-infecting haemopoietic stem cell (HSC)-enriched cells, derived from wild type mice, with a dominant negative protein kinase C alpha (PKCa-KR) construct and culturing the cells either in in vitro or in vivo B cell generation systems. This resulted in the spontaneous generation of B lineage cells that resemble human CLL cells both at the phenotypic and molecular level, being refractory to apoptosis - possibly due to an upregulation in Bcl-2 and Mcl-1 expression levels, and arrested in G0/G1 ex vivo (Cancer Res. 66:527). These studies indicated that subversion of PKCa activity served to initiate the development of CLL. In this application, we show preliminary data demonstrating that our mouse CLL cells can be generated from recombinase activating gene deficient mice, which exhibit a development block at an early stage of B cell development, thus establishing that CLL can be derived from either an HSC or an early B cell progenitor population. Therefore, this proposal will phenotypically identify and molecularly characterise the CLL cell of origin both in our novel CLL mouse model and importantly in samples derived from CLL patients. We will then functionally characterise the mouse and human leukaemia initiating cell, both in vitro and in vivo. Collectively, the proposed studies will provide a deeper understanding of the biology of CLL, which may in turn illuminate novel avenues for therapeutic intervention.