Nicola Foot

Bortezomib, doxorubicin and dexamethasone (PAD) front‐line treatment of multiple myeloma: updated results after long‐term follow‐up

Bortezomib, doxorubicin and dexamethasone (PAD) was evaluated as induction before stem cell transplantation in newly diagnosed multiple myeloma (MM) patients, using bortezomib 1·3 mg/m2 (PAD1, N = 21) or 1·0 mg/m2 (PAD2, N = 20). Complete/very good partial response rates with PAD1/PAD2 were 62%/42% postinduction and 81%/53% post‐transplant. Progression‐free survival (29 vs. 24 months), time to re‐treatment (36 vs. 29 months) and overall survival (1 year: 100% vs. 95%; 2 years: 95% vs. 73%) were statistically similar but favoured PAD1 versus PAD2. Toxicity was lower in PAD2; bortezomib dose reduction may help manage toxicities while retaining efficacy. PAD is highly active as front‐line induction in MM.

Detection of Chromosome Abnormalities Pre–High-Dose Treatment in Patients Developing Therapy-Related Myelodysplasia and Secondary Acute Myelogenous Leukemia After Treatment for Non-Hodgkin’s Lymphoma

PURPOSE To assess whether pre-high-dose therapy (HDT)-related factors play a critical role in the development of therapy-related myelodysplasia (tMDS) or secondary acute myelogenous leukemia (sAML). PATIENTS AND METHODS Twenty-nine of 230 patients with a primary diagnosis of non-Hodgkins lymphoma (NHL) developed tMDS/sAML after HDT comprising cyclophosphamide and total-body irradiation (TBI) supported by autologous hematopoietic progenitor cells. G-banding and fluorescence in-situ hybridization (FISH) were used to detect clonal cytogenetic abnormalities. RESULTS The majority of patients showed complex karyotypes at diagnosis of tMDS/sAML containing, in particular, complete or partial loss of chromosomes 5 and/or 7. Using single locus-specific FISH probes, significant levels of clonally abnormal cells were found before HDT in 20 of 20 tMDS/sAML patients screened, compared with three of 24 patients screened who currently have not developed tMDS/sAML, at a median follow-up of 5.9 years after HDT. CONCLUSION Prior cytotoxic therapy may play an important etiologic role and may predispose to the development of tMDS/sAML. Using a triple FISH assay designed to detect loss of chromosomal material from 5q31, 7q22, or 13q14, significant levels of abnormal cells can be detected before HDT and may predict which patients are at increased risk of developing secondary disease. Further prospective evaluation of this FISH assay is warranted to determine its predictive power in this setting.

Localisation of a novel region of recurrent amplification in follicular lymphoma to an ∼6.8 Mb region of 13q32‐33

Follicular lymphoma (FL) is characterised by the presence of the t(14;18)(q32;q21) and represents ∼25% of new cases of non‐Hodgkins lymphoma. While the t(14;18) is a well‐documented rearrangement, the role of secondary cytogenetic abnormalities in the development and progression of these tumours remains unclear. Comparative genomic hybridisation was used to characterise changes in DNA copy number in tumour DNA from patients with this malignancy. The mean numbers of deletion and amplification events found in each of the 45 samples studied were 1.8 and 2.3, respectively. Regions of recurrent (>10% tumour samples) gain involved chromosomes 2p13‐16 (16%), 7 (20%), 12 (16%), 13q21‐33 (18%), 18 (27%), and X (36%) and frequent losses localised to 6q (29%) and 17p (20%). Amplification of chromosome 13 represents a novel finding in FL. The minimal amplified region was refined to a 6.8‐Mb interval of 13q32‐33 between the BAC clones 88K16 and 44H20 by fluorescence in situ hybridisation studies using metaphase chromosomes derived from tumour material. There are a number of reports in the literature suggesting that amplification of chromosome 13 also occurs in other human cancers. The location of the putative oncogene on 13q described here in follicular and transformed lymphoma may also be important in the evolution of many other malignancies.

Bortezomib, low‐dose intravenous melphalan, and dexamethasone for patients with relapsed multiple myeloma

This multicenter phase I/II study investigated the maximum tolerated dose (MTD), safety, and efficacy of low dose intravenous (IV) melphalan in combination with bortezomib for patients with relapsed multiple myeloma (MM). Patients received bortezomib 1·3 mg/m2 on days 1, 4, 8, and 11 and escalating doses of IV melphalan (2·5–10·0 mg/m2) on day 2 of a 28‐day cycle for a maximum of eight cycles. Dexamethasone 20 mg was added for progressive or stable disease. Fifty‐three patients were enrolled. The MTD was defined at melphalan 7·5 mg/m2 and bortezomib 1·3 mg/m2. The overall response rate (ORR) was 68% (23% complete or near‐complete responses [CR/nCR]) whilst at the MTD (n = 33) the ORR was 76% (34% CR/nCR). After median follow‐up of 17 months, the median progression free survival was 10 months, rising to 12 months at the MTD (P < 0·05 vs. non‐MTD regimens). The median overall survival was 28 months, but was not yet reached at the MTD. Grade 3/4 adverse events included thrombocytopenia (62%), neutropenia (57%), infection (21%), and neuropathy (15%). Bortezomib and low‐dose IV melphalan combination therapy is a safe and highly effective regimen for patients with relapsed MM. These data suggest further investigation of this combination is warranted.

Prospective gene expression analysis accurately subtypes acute leukaemia in children and establishes a commonality between hyperdiploidy and t(12;21) in acute lymphoblastic leukaemia

We have prospectively analysed and correlated the gene expression profiles of children presenting with acute leukaemia to the Royal London and Great Ormond Street Hospitals with morphological diagnosis, immunophenotype and karyotype. Total RNA extracted from freshly sorted blast cells was obtained from 84 lymphoblastic [acute lymphoblastic leukaemia (ALL)], 20 myeloid [acute myeloid leukaemia (AML)] and three unclassified acute leukaemias and hybridised to the high density Affymetrix U133A oligonucleotide array. Analysis of variance and significance analysis of microarrays was used to identify discriminatory genes. A novel 50‐gene set accurately identified all patients with ALL and AML and predicted for a diagnosis of AML in three patients with unclassified acute leukaemia. A unique gene set was derived for each of eight subtypes of acute leukaemia within our data set. A common profile for children with ALL with an ETV6–RUNX1 fusion, amplification or deletion of ETV6, amplification of RUNX1 or hyperdiploidy with an additional chromosome 21 was identified. This suggests that these rearrangements share a commonality in biological pathways that maintains the leukaemic state. The gene TERF2 was most highly expressed in this group of patients. Our analyses demonstrate that not only is microarray analysis the single most effective tool for the diagnosis of acute leukaemias of childhood but it has the ability to identify unique biological pathways. To further evaluate its prognostic value it needs to be incorporated into the routine diagnostic analysis for large‐scale clinical trials in childhood acute leukaemias.

Expression profile of wild-type ETV6 in childhood acute leukaemia

Summary. Comparative expression analysis of wild‐typeETV6 in the disease state showed an absence of expression in ETV6–CBFA2 acute lymphoblastic leukaemia (ALL) when compared with non‐ETV6–CBFA2 ALL and acute myeloid leukaemia. Fluorescent in‐situ hybridization and loss of heterozygosity studies showed that 73% of the ETV6–CBFA2 samples had a fully or partially deleted second ETV6 allele, explaining the lack of wild‐type expression in these patients. Although the second ETV6 allele was identified in the remaining patients, no ETV6 expression was detected. These observations support the hypothesis that loss of ETV6 expression is a critical secondary event for leukaemogenesis in ETV6–CBFA2 ALL.

Assignment1 of brain acid-soluble protein 1 (BASP1) to human chromosome 5p15.1→p14, differential expression in human cancer cell lines as a result of alterations in gene dosage

Brain acid-soluble protein 1 (BASP1, previously Neuronal Tissue Enriched Acidic protein NAP-22 Unigene Hs. 79516) (Park et al., 1988; Maekawa et al., 1999) encodes a neuronenriched Ca(2+)-dependent calmodulin-binding protein of unknown function. An Expressed Sequence Tag (EST) corresponding to BASP1 was found by Northern analysis to be differentially expressed in a panel of human cancer cell lines with highest expression in the cervical adenocarcinoma cell line, HeLa S3. In evaluating the role of BASP1 in cancer the gene was assigned to human chromosome 5p15.1→p14 by fluorescence in situ hybridisation (FISH) and its DNA copy number in these cell lines determined. Materials and methods

Assignment1 of B-cell lymphoma 6, member B (zinc finger protein) gene (BCL6B) to human chromosome 17p13.1 by in situ hybridization

Rearrangement of the B cell lymphoma 6 (BCL6) gene on chromosome 3q27 is a recurrent translocation event in nonHodgkin’s lymphoma (Bastard et al., 1994; Chaganti et al., 1998). The frequency of these rearrangements depend on the histology of the tumour and are found in approximately 10% and 30% of Follicular and Diffuse large B cell lymphomas respectively (Ohno and Fukuhara, 1997). BCL6B (BCL6 member B, previously BAZF), a novel BCL6 homologue (Okabe et al., 1998), has recently been cloned in the mouse. There are three regions of homology between the two proteins, the BTB/ POZ domain at the NH2-terminal region, a perfectly conserved 17 aa sequence in the middle region and the zinc finger domain at the COOH-terminus. All three domains are required for BCL6B to bind the BCL6 DNA binding sequence and act as a transcriptional repressor, suggesting that both proteins regulate similar target genes. This raises the possibility that BCL6B may also be involved in lymphomagenesis. Materials and methods