Genetic Defects

Genetic aberrations in CLL are significantly associated with clinical outcomes. These include deletions 13q14, 11q22-q23 and 17p13, trisomy 12, and more recently due to next generation sequencing, mutations in the tumour suppressor gene TP53, NOTCH1, SF3B1, BIRC3, MYD88 and Wnt signalling pathway genes have also been determined. Identification of such genetic defects are useful in gaining further understanding of the pathways involved in CLL pathogenesis (Nadeu et al., 2016; Ten Hacken & Burger, 2014; Zhang & Kipps, 2014). Some mutations are seen in all CLL cells from a subclonal population, whereas other mutations may be specific to only a subset of cells (Zhang & Kipps, 2014). Furthermore, these genetic mutations are more prevalent in chemotherapy-refractory patients and those with Richter’s transformation, which suggests that they are gained during disease progression (Efremov & Laurenti, 2014).

The evolution of mutations has been studied in 149 patients with CLL using whole-exome sequencing and copy number to determine the proportion of cancer cells containing each mutation. Mutations were found to be either mainly clonal (e.g. trisomy 12, del 13q and MYD88) associated with early events in CLL, or predominantly subclonal (e.g. TP53, SF3B1) associated with later events in CLL. Subsequently, a sample group of 18 patients were analysed at two time points: of 12 patients treated with chemotherapy 10 underwent clonal evolution mostly involving subclones with TP53 or SF3B1 mutations, whereas only 1 out of 6 patients not receiving treatment had clonal evolution. In addition, the subclonal mutations were determined to be an independent risk factor for rapid disease progression (Landau et al., 2013).

Next-generation sequencing (NGS), as well as identifying the above mentioned genetic defects, has also revealed a number of other novel, recurrent mutations in CLL including KLH6, POT1, and XPO1, although the possible clinical significance of these mutations is yet to be determined (Sutton et al., 2015). Targeted NGS has been demonstrated to provide reproducible and accurate identification of mutations in CLL. In a study of 188 patients with CLL, targeted NGS showed that 63% of patients carried at least one mutation with 84% of these being mutations in ATM, BIRC3, NOTCH1, SF3B1 and TP53 genes. In the future, this technique could potentially be used for routine screening in the clinic without the need for Sanger sequencing but as yet, it requires further validation (Sutton et al., 2015).

Whole-genome and exome sequencing may be a potential useful tool in the identification of clinically relevant mutations in patients with CLL. In one study, this whole-genome sequencing was used to identify the two major subtypes of CLL, namely CLL with unmutated immunoglobulin genes (unmutated IGHV) and CLL with mutated immunoglobulin genes (mutated IGHV). Combined with exome sequencing, 46 somatic mutations were detected encoding proteins potentially affecting gene function. Further analysis of data from a larger cohort of patients with CLL identified NOTCH1, XPO1, MYD88 and KLHL genes that were recurrently mutated. NOTCH1 and XPO1 mutations were mainly associated with unmutated IGHV CLL whereas MYD88 and KLHL mutations tended to be associated with mutated IGHV CLL. Combined with further data, it was strongly indicated that NOTCH1, MYD88 and XPO1 mutations contribute to the evolution of CLL (Puente et al., 2011). A more recent whole-exome sequence analysis of 538 CLL and matched germline DNA samples identified 44 recurrently mutated genes and 11 recurrent somatic copy number variations including previously unrecognized cancer drivers (RPS15, IKZF3) (Landau et al., 2015).

A brief outline of characterised genetic defects in relation to the pathology of CLL is given below and their clinical impact is further discussed in the Prognosis section.

  • TP53: Somatic mutations in the tumour suppressor gene TP53 occur in around 10% of CLL at diagnosis (Rossi et al., 2009) and 4–37% of all patients with CLL (Hallek, 2017). TP53 aberrations confer very poor prognosis (Hallek, 2017) and are associated with fludarabine-refractory CLL and CLL with prolymphocytic morphology (Rossi et al., 2009). Such mutations also confer short survival and chemorefractoriness, independent of del 17p13 (Rossi et al., 2009; Stilgenbauer et al., 2014).
  • Del 17p: Deletions in chromosome 17 (del 17p), and most commonly del 17p13 which results in loss of the TP53 tumour suppressor gene, are observed in 5–8% of CLL cases at diagnosis (Efremov & Laurenti, 2014; Hallek, 2015;). Del 17p prognosticates poor outcome and chemorefractoriness (Byrd et al., 2014; Döhner et al., 2000; Hallek, 2017; Rossi et al., 2009; Stilgenbauer et al., 2014).
  • Del 13q14: This is the most common genetic aberration seen in CLL occurring in 50–60% of patients. 13q14 deletion disrupts the expression of several anti-apoptotic and cell cycle regulatory proteins (Efremov & Laurenti, 2014).
  • Del 11q: Approximately 25% of patients with advanced stage disease and 10% patients with early stage disease have deletions in chromosome 11 which confers bulky lymphadenopathy, rapid progression and poor survival (Hallek, 2017). Specifically, del 11q22-23 results in loss of the ATM gene (encodes DNA damage response kinase), seen in 18% of cases at diagnosis (Byrd et al., 2014; Efremov & Laurenti, 2014; Stilgenbauer et al., 2014).
  • Trisomy 12: An extra copy of chromosome 12 is observed in 10–20% of CLL cases at diagnosis. Currently, little is known about the genes involved in CLL pathogenesis as a result of trisomy 12 (Efremov & Laurenti, 2014; Hallek, 2017).
  • NOTCH1: Encodes the NOTCH1 transcription factor which is involved in regulation of pathways that induce differentiation of haematopoietic progenitor cells into T cells, and development of mature B cells into antibody-secreting cells (Zhang & Kipps, 2014). NOTCH1 mutations occur in approximately 10% of de novo CLL and 15–20% of relapsed/progressive CLL, and are associated with high-risk CLL (Rossi et al., 2009; Zhang & Kipps, 2014).
  • SF3B1: Encodes a splicing factor sub-unit (SF3B1), a component of spliceosomes which are involved in pre-mRNA processing, specifically in genes controlling cell cycle progression and apoptosis. Around 10% of newly diagnosed CLL and 17% of progressive late-stage CLL have mutations in SF3B1 which may promote cell proliferation and/or survival.
  • BIRC3: Encodes an inhibitor of apoptosis (IAP) protein (BIRC3). CLL cells with mutations in BIRC3 result in NF-κB activation and are less responsive to conventional chemotherapy compared to CLL cells without BIRC3 mutations (Zhang & Kipps, 2014). 
  • MYD88: Mutations in MYD88, which encodes an adaptor molecule of the Toll-like receptor (TLR) complex, are observed in 3–10% of newly diagnosed CLL (Zhang & Kipps, 2014).
  • Wnt signalling pathway genes: Mutations in genes encoding Wnt signalling proteins are prevalent in CLL. Wnt signalling is activated in CLL, especially in disease with unmutated IGHV. The aberrant activation of this pathway may therefore be due to the accumulation of genetic mutations seen in Wnt genes (Zhang & Kipps, 2014).