Ordering of mutations in acute myeloid leukemia with partial tandem duplication of MLL (MLL-PTD).

Partial tandem duplication of MLL (MLL-PTD) characterizes acute myeloid leukemia (AML) patients often with a poor prognosis. To understand the order of occurrence of MLL-PTD in relation to other major AML mutations and to identify novel mutations that may be present in this unique AML molecular subtype, exome and targeted sequencing was performed on 85 MLL-PTD AML samples using HiSeq-2000. Genes involved in the cohesin complex (STAG2), a splicing factor (U2AF1) and a poorly studied gene, MGA were recurrently mutated, whereas NPM1, one of the most frequently mutated AML gene, was not mutated in MLL-PTD patients. Interestingly, clonality analysis suggests that IDH2/1, DNMT3A, U2AF1 and TET2 mutations are clonal and occur early, and MLL-PTD likely arises after these initial mutations. Conversely, proliferative mutations (FLT3, RAS), typically appear later, are largely subclonal and tend to be unstable. This study provides important insights for understanding the relative importance of different mutations for defining a targeted therapeutic strategy for MLL-PTD AML patients.

Host genetic variants of ABCB1 and IL15 influence treatment outcome in paediatric acute lymphoblastic leukaemia.

BACKGROUND: Host germline variations and their potential prognostic importance is an emerging area of interest in paediatric ALL. METHODS: We investigated the associations between 20 germline variations and various clinical end points in 463 children with ALL. RESULTS: After adjusting for known prognostic factors, variants in two genes were found to be independently associated with poorer EFS: ABCB1 T/T at either 2677 (rs2032582) or 3435 (rs1045642) position (P=0.003) and IL15 67276493G/G (rs17015014; P=0.022). These variants showed a strong additive effect affecting outcome (P<0.001), whereby patients with both risk genotypes had the worst EFS (P=0.001), even after adjusting for MRD levels at the end of remission induction. The adverse effect of ABCB1 T/T genotypes was most pronounced in patients with favourable cytogenetics (P=0.011) while the IL15 67276493G/G genotype mainly affected patients without common chromosomal abnormalities (P=0.022). In both cytogenetic subgroups, increasing number of such risk genotypes still predicted worsening outcome (P<0.001 and=0.009, respectively). CONCLUSION: These results point to the prognostic importance of host genetic variants, although the specific mechanisms remain unclarified. Inclusion of ABCB1 and IL15 variants may help improve risk assignment strategies in paediatric ALL.

Hot and steamy: outbreak of Bacillus cereus in Singapore associated with construction work and laundry practices.

BACKGROUND: A sudden increase in invasive infections caused by Bacillus cereus group organisms prompted an investigation at the National University Hospital in Singapore. AIM: To describe the investigation and management and subsequent difficulties controlling the outbreak. METHODS: Clinical case reviews were performed on all patients with B. cereus group recovered from clinical samples. Widespread environmental sampling was performed followed by review of hospital ventilation systems, domestic cleaning and laundry practices. FINDINGS: B. cereus was recovered from 171 patients during a six-month period coinciding with large-scale construction work beside the hospital. Most patients presented with bacteraemia (146/171; 85.4%) with 46/171 (26.9%) requiring extended treatment courses with vancomycin or other interventions. Sampling confirmed extensive airborne dispersal inside the hospital, including isolation rooms and air-conditioned wards. Hospital linen was heavily contaminated [7403 cfu/cm(2); 95% confidence interval (CI): 6349-8457; for 30 towels sampled], encouraged by inappropriate storage in airtight plastic bags (4437 cfu/cm(2); CI: 3125-5750) compared with storage in porous canvas bags (166 cfu/cm(2); CI: 76-256; P < 0.001). Interventions introduced included revision of laundry practices, transport and storage of hospital linen and towels; bleach-based environmental cleaning; and upgrading of ventilation systems throughout the hospital. Clinical case numbers returned to baseline levels within three months, only to rise again following relaxation of laundry practices. CONCLUSIONS: Construction work beside this Singapore hospital encouraged heavy contamination of air and environment with Bacillus spp., assumed to be responsible for the outbreak described. Failure to maintain revised laundry practices allowed resurgence of clinical cases, particularly among immunocompromised patients.

Novel HLA class I and II alleles identified during routine registry typing in 2010.

Twenty-one novel human leukocyte antigen (HLA) class I and class II alleles are described: seven HLA-A alleles, five HLA-C alleles, five HLA-B alleles and four HLA-DRB1 alleles. Seventeen (∼81%) of the 21 novel alleles are single nucleotide substitution variants when compared with their most homologous allele. Nine of these single nucleotide variants cause an amino acid substitution, while eight are silent substitutions. The remaining novel alleles differ from their most similar allele by two to three nucleotide substitutions. One novel HLA-C locus allele encodes an amino acid change at codon 10 that previously has not been reported to be polymorphic for this locus. Some of the new alleles encode novel codons and unique amino acid changes at polymorphic positions in the HLA-A (codons 24 and 156) and HLA-B (codons 40 and 115) loci.

Dedicated cytogenetics factor is critical for improving karyotyping results for childhood leukaemias - experience in the National University Hospital, Singapore 1989-2006.

INTRODUCTION: Childhood leukaemia accounts for more than 40% of new childhood cancer cases. Karyotyping of cytogenetic abnormalities in such cases continues to provide critical prognostic information which allows the delivery of an appropriate intensity of treatment. Unfortunately, karyotyping of childhood leukaemia is difficult, laborious and often unsuccessful. Banding resolution tends to be poor unlike routine antenatal cytogenetics. The aim of the study is to highlight the benefit of dedicated cytogenetics in improving karyotyping results. MATERIALS AND METHODS: We analysed the impact of setting up a team of cytogeneticists in the National University Hospital (NUH) on the success of karyotyping, evaluating cytogenetic data collected from 1989 to 2006. From 1989 to 2006, 4789 cases have been processed. Among them, 369 newly diagnosed and relapsed childhood acute leukaemia cases [281 acute lymphoblastic leukaemia (ALL) and 88 acute myeloid leukaemia (AML)] have been diagnosed at NUH. A dedicated cytogenetics laboratory with clearly defined standard operating procedures and quality control was set up in 2002. It used the established recommendation of a complete analysis of at least 20 metaphases per analysis. RESULTS: Overall, the frequency of successful karyotyping was significantly higher (P = 0.002) at 90.7% (185/204) from 2002-2006 compared to 79.4% (131/165) from 1989-2001. For ALL cases, the success rate improved from 77.6% (97/125) in 1989 to 2001 to 89.1% (139/156) in the 2002 to 2006 cohort. For AML, the success rate also was significantly improved (P = 0.04) from 85% (34/40) to 95.8% (46/48). Significantly, this high rate of success is still maintained despite a yearly increase in volume. CONCLUSION: The establishment of a dedicated cytogenetics service leads to an improvement in results.

Characterization of new HLA-B and -DRB1 alleles from Singapore.

Six novel alleles, three human leukocyte antigen (HLA)-B and three HLA-DRB1 alleles, are described. Five of the variants are single nucleotide substitutions from their most homologous allele, of which three result in amino acid changes (B*3572, *9509 and DRB1*1157) and two are silent substitutions (B*370103 and DRB1*150204). DRB1*0462 differs by three nucleotide substitutions that alter two amino acids.

Identification of nine new HLA class I alleles in volunteers from the Singapore stem cell donor registries.

Based on unusual probe hybridization patterns, two human leukocyte antigen (HLA)-A, five HLA-B, and two HLA-C alleles (A*240208, A*3211Q; B*1822, B*3938Q, B*4606, B*4607N, B*5139; Cw*0321, Cw*0734) were identified in individuals from the Singapore Bone Marrow Donor Program and Singapore Cord Blood Bank. Eight of the nine alleles encode amino acid substitutions altering the antigen-binding region including three alleles with changes altering a cysteine at codon 164 (A*3211Q, B*3938Q, B*4607N). This substitution either eliminates a key disulfide bond or results in a stop codon, both likely affecting the expression of the HLA molecules. Only one allele (A*240208) carries a synonymous substitution.

Thiopurine methyltransferase polymorphisms in a multiracial asian population and children with acute lymphoblastic leukemia.

The purpose of this study was to determine the frequency of thiopurine methyltransferase (TPMT) polymorphisms in a multiracial Asian population and to assess its relevance in the management of childhood acute lymphoblastic leukemia (ALL). Six hundred unrelated cord blood samples from 200 Chinese, Malay, and Indian healthy newborns were collected at the National University Hospital, Singapore; an additional 100 children with ALL were analyzed for five of the commonly reported TPMT variant alleles using polymerase chain reaction/restriction fragment length polymorphism and allele-specific polymerase chain reaction-based assays. In the cord blood study, the TPMT*3C variant was detected in all three ethnic groups; Chinese, Malays, and Indians had allele frequencies of 3%, 2.3%, and 0.8%, respectively. The TPMT*3A variant was found only among the Indians at a low allele frequency of 0.5%. The TPMT*6 variant was found in one Malay sample. Among the children with ALL, two white and one Chinese were heterozygous for the TPMT*3A variant and showed intermediate sensitivity to 6-mercaptopurine during maintenance therapy. Three Chinese patients and one Malay patient were heterozygous for the TPMT*3C variant. Mercaptopurine sensitivity could be validated in only one out of four TPMT*3C heterozygous patients. The overall allele frequency of the TPMT variants in this multiracial population was 2.5%. The TPMT*3C was the most common variant allele; TPMT*3A and TPMT*6 were rare. These results support the feasibility of performing TPMT genotyping in all children diagnosed with acute leukemia to minimize toxicity from thiopurine chemotherapy.