Faith E. Davies

A novel prognostic model in myeloma based on co-segregating adverse FISH lesions and the ISS: analysis of patients treated in the MRC Myeloma IX trial.

The association of genetic lesions detected by fluorescence in situ hybridization (FISH) with survival was analyzed in 1069 patients with newly presenting myeloma treated in the Medical Research Council Myeloma IX trial, with the aim of identifying patients associated with the worst prognosis. A comprehensive FISH panel was performed, and the lesions associated with short progression-free survival and overall survival (OS) in multivariate analysis were +1q21, del(17p13) and an adverse immunoglobulin heavy chain gene (IGH) translocation group incorporating t(4;14), t(14;16) and t(14;20). These lesions frequently co-segregated, and there was an association between the accumulation of these adverse FISH lesions and a progressive impairment of survival. This observation was used to define a series of risk groups based on number of adverse lesions. Taking this approach, we defined a favorable risk group by the absence of adverse genetic lesions, an intermediate group with one adverse lesion and a high-risk group defined by the co-segregation of >1 adverse lesion. This genetic grouping was independent of the International Staging System (ISS) and so was integrated with the ISS to identify an ultra-high-risk group defined by ISS II or III and >1 adverse lesion. This group constituted 13.8% of patients and was associated with a median OS of 19.4 months.

Prediction of outcome in newly diagnosed myeloma: a meta-analysis of the molecular profiles of 1905 trial patients

Robust establishment of survival in multiple myeloma (MM) and its relationship to recurrent genetic aberrations is required as outcomes are variable despite apparent similar staging. We assayed copy number alterations (CNA) and translocations in 1036 patients from the NCRI Myeloma XI trial and linked these to overall survival (OS) and progression-free survival. Through a meta-anlysis of these data with data from MRC Myeloma IX trial, totalling 1905 newly diagnosed MM patients (NDMM), we confirm the association of t(4;14), t(14;16), t(14;20), del(17p) and gain(1q21) with poor prognosis with hazard ratios (HRs) for OS of 1.60 (P=4.77 × 10−7), 1.74 (P=0.0005), 1.90 (P=0.0089), 2.10 (P=8.86 × 10−14) and 1.68 (P=2.18 × 10−14), respectively. Patients with ‘double-hit’ defined by co-occurrence of at least two adverse lesions have an especially poor prognosis with HRs for OS of 2.67 (P=8.13 × 10−27) for all patients and 3.19 (P=1.23 × 10−18) for intensively treated patients. Using comprehensive CNA and translocation profiling in Myeloma XI we also demonstrate a strong association between t(4;14) and BIRC2/BIRC3 deletion (P=8.7 × 10−15), including homozygous deletion. Finally, we define distinct sub-groups of hyperdiploid MM, with either gain(1q21) and CCND2 overexpression (P<0.0001) or gain(11q25) and CCND1 overexpression (P<0.0001). Profiling multiple genetic lesions can identify MM patients likely to relapse early allowing stratification of treatment.

Assessing the effect of obesity-related traits on multiple myeloma using a Mendelian randomisation approach

Multiple myeloma (MM) accounts for around 15% of new cases and 20% of deaths amongst patients diagnosed with haematological malignancy. To date, few risk factors have been robustly confirmed for MM, these include increasing age, male sex, black race and a family history of MM.1

Patients in the MRC Myeloma XI trial receiving induction chemotherapy for newly diagnosed multiple myeloma containing dexamethasone at full or moderated dose have significantly reduced half-life of IgG

Chronic myeloid leukaemia (CML) is characterised by the presence of the fusion protein BCR-ABL1. The addiction of CML cells to the tyrosine kinase (TK) activity of the oncoprotein has been successfully exploited by the introduction of tyrosine kinase inhibitors (TKI), such as imatinib, which have shown a great success at managing the disease. However, these compounds fail to eradicate a primitive cell population, the leukaemic stem cells (LSCs), which persist in the patients. This translates in the need of life-long therapy for most of the patients, meaning a higher risk of treatment side effects and the prolonged psychological burden of living a leukaemia patient. Life-long therapy is also translating in a continuous increase in CML prevalence in developed countries and sustaining a big patient population on TKI treatment is becoming a challenge for national health systems. Recent reports in chronic myeloid leukaemia biology have confirmed that CML LSCs are not addicted to the TK activity of BCR-ABL1 and they retain repopulation and leukaemic properties even during BCR-ABL1 TK inhibition. Thus, the discovery of new therapeutic targets capable of eliminating this cell population is required for curing the disease. Previous reports have already shown great success at reducing the number of CML LSCs by targeting JAK2, STAT5, EZH2, MYC and p53 pathways as well as autophagocytosis. However, none of them have shown complete eradication of the clone and they failed to define a global gene expression signature that may explain the persistence of CML LSCs during TKI treatment. Taken together, the work presented in this thesis confirms the existence of a BCR-ABL1 transcriptional signature in CML LSCs. Also, it shows that targeting CD33, a member of the TKIi signature, reduces the number of CML CD34+ cells and induces a transcriptional and phenotypic change towards a cycling and repopulating cell population. Additionally, the use of the TKIi signature has shown potential as a molecular biomarker for predicting TKI response in CML patients.

B-cell malignancies: capture-sequencing strategies for identification of gene rearrangements and translocations into immunoglobulin gene loci

Aberrant chromosomal translocations involving the immunoglobulin heavy chain (IGH) locus and clonal rearrangements of the variable (V), diversity (D), joining (J) segments (V(D)J) of the immunoglobulin gene are implicated in the initiation of mature B-cell malignan- cies, including myeloma. Analysis of these events provides information useful for diagnosis and prediction of prognosis, and also provides a useful monitoring strategy for response to treatment in patients with these diseases. Current methods for identification of these events are not gener - ally applicable and give a biased picture of the prognostic significance and clinical relevance of these events. Novel methodologies based on next-generation sequencing are a new and more efficient genetic tool that can be used to screen and characterize these changes at the nucleotide level, offering a deeper and better understanding of the genetic basis of these complex diseases. In this work, we provide a review of the molecular basis of B-cell neoplasms, the methods used to detect them, and how they translate into the clinic.

Deletion of 16q identified by FISH is an independent adverse prognostic marker in multiple myeloma

Chromosomal translocations lead to oncogene activation in a significant number of haematological malignancies. Those involving the immunoglobulin heavy chain locus, IGH, at chromosome band 14q32 are frequently observed in B-cell malignant proliferation. A small number have been described in B-cell precursor acute lymphoblastic leukaemia (BCP-ALL). However, their biological and clinical significance is currently unknown. Detailed fluorescence in situ hybridisation (FISH) and molecular studies were carried out on a series of BCP-ALL patients with chromosomal abnormalities involving 14q32. Novel and recurrent translocations affecting different chromosomes were highlighted. Refined FISH mapping identified putative IGH partner genes at, or flanking, the translocation breakpoints. Four translocations: two previously reported, t(14;19)(q32;q13), t(8;14)(q11;q32), and two novel, t(14;14)(q11;q32)/ inv(14)(q11q32) and t(14;20)(q32;q13), were identified. Molecular analyses showed that four different members of the CAATT enhancer binding protein (CEBP) gene family were involved: CEBPA (19q13, n59), CEBPD (8q11, n58), CEBPE (14q11, n53) and CEBPB (20q13, n52). One patient with a t(14;19)(q32;q13) was observed to involve the fifth family member CEBPG (19q13, n51). Breakpoints were located within the 30 untranslated region (UTR) of CEBPA and either 30 UTR or 50 of CEBPE, whereas breakpoints in 8q11 were B30 kb centromeric of CEBPD. Where material was available, over-expression of target genes was shown by quantitative real-time PCR. Overall, this study has demonstrated for the first time the involvement of five members of the same gene family in a single subtype of haematological disease. It has indicated that transcriptional upregulation of CEBP gene family members, by juxtaposition to IGH, is important in BCP-ALL: a mechanism in complete contrast to that involving CEPBA in acute myeloid leukaemia.