Infant acute lymphoblastic leukemia (ALL) with MLL rearrangements (MLL-R) represents a distinct leukemia with a poor prognosis. To define its mutational landscape, we performed whole genome, exome, RNA and targeted DNA sequencing on 65 infants (47 MLL-R and 18 non-MLL-R) and 20 older children (MLL-R cases) with leukemia. Our data demonstrated infant MLL-R ALL to have one of the lowest frequencies of somatic mutations of any sequenced cancer, with the predominant leukemic clone carrying a mean of 1.3 non-silent mutations. Despite the paucity of mutations, activating mutations in kinase/PI3K/RAS signaling pathways were detected in 47%. Surprisingly, however, these mutations were often sub-clonal and frequently lost at relapse. In contrast to infant cases, MLL-R leukemia in older children had more somatic mutations (a mean of 6.5/case versus 1.3/case, P=7.15×10−5) and contained frequent mutations (45%) in epigenetic regulators, a category of genes that with the exception of MLL was rarely mutated in infant MLL-R ALL.

SUMMARY To define the mutation spectrum in non-Down syndrome acute megkaryoblastic leukemia (non-DS-AMKL), we performed transcriptome sequencing on diagnostic blasts from 14 pediatric patients and validated our findings in a recurrency/validation cohort consisting of 34 pediatric and 28 adult AMKL leukemia samples. Our analysis identified a cryptic chromosome 16 inversion [inv(16)(p13.3q24.3)] in 27% of pediatric cases, which encodes a CBFA2T3-GLIS2 fusion protein. Expression of CBFA2T3-GLIS2 in Drosophila and murine hematopoietic cells induced bone morphogenic protein (BMP) signaling, and resulted in a marked increase in the self-renewal capacity of hematopoietic progenitors. These data suggest that expression of CBFA2T3-GLIS2 directly contributes to leukemogenesis.

Monotherapy with (-)2',3'-dideoxy-3'-thiacytidine (3TC) leads to the appearance of a drug-resistant variant of human immunodeficiency virus-type 1 (HIV-1) with the methionine-184 --> valine (M184V) substitution in the reverse transcriptase (RT). Despite resulting drug resistance, treatment for more than 48 weeks is associated with a lower plasma viral burden than that at baseline. Studies to investigate this apparent contradiction revealed the following. (i) Titers of HIV-neutralizing antibodies remained stable in 3TC-treated individuals in contrast to rapid declines in those treated with azidothymidine (AZT). (ii) Unlike wild-type HIV, growth of M184V HIV in cell culture in the presence of d4T, AZT, Nevirapine, Delavirdine, or Saquinavir did not select for variants displaying drug resistance. (iii) There was an increase in fidelity of nucleotide insertion by the M184V mutant compared with wild-type enzyme.

Mice carrying the Tight skin (Tsk) mutation have thickened skin and visceral fibrosis resulting from an accumulation of extracellular matrix molecules. These and other connective tissue abnormalities have made Tskl + mice models for scleroderma, hereditary emphysema, and myocardial hypertrophy. Previously we localized Tsk to mouse chromosome 2 in a region syntenic with human chromosome 15. The microfibrillar glycoprotein gene, fibrillin 1 (FBN1), on human chromosome 15q, provided a candidate for the Tsk mutation. We now demonstrate that the Tsk chromosome harbors a 30- to 40-kb genomic duplication within the Fbn1 gene that results in a larger than normal in-frame Fbn1 transcript. These findings provide hypotheses to explain some of the phenotypic characteristics of Tskl + mice and the lethality of Tsk/Tsk embryos.

BackgroundNeuroblastomas are characterized by hemizygous 1p deletions, suggesting that a tumor suppressor gene resides in this region. We previously mapped the smallest region of consistent deletion to a 2-Mb region of 1p36.31 that encodes 23 genes. Based on mutation analysis, expression pattern, and putative function, we identified CHD5 as the best tumor suppressor gene candidate.MethodsWe determined the methylation status of the CHD5 gene promoter in NLF and IMR5 (with 1p deletion) and SK-N-SH and SK-N-FI neuroblastoma cell lines using methylation-specific sequencing and measured CHD5 mRNA expression by reverse transcription polymerase chain reaction in cells treated with or without 5-aza-2-deoxycytidine, an inhibitor of DNA methylation. We transfected the cells with CHD5 and antisense (AS) CHD5 DNA to assess the effect of CHD5 overexpression and suppression, respectively, on colony formation in soft agar and growth of xenograft tumors in athymic mice. We also analyzed the association of CDH5 expression with outcomes of 99 neuroblastoma patients. Statistical tests were two-sided.ResultsCHD5 expression was very low or absent in neuroblastoma cell lines. The CHD5 promoter was highly methylated in NLF and IMR5 lines, and CHD5 expression increased after treatment with 5-aza-2-deoxycytidine. Clonogenicity and tumor growth were abrogated in NLF and IMR5 cells overexpressing CHD5 compared with antisense CHD5 (clonogenicity: mean no. of colonies per plate, NLF-CHD5, 43 colonies, 95% confidence interval [CI] = 35 to 51 colonies, vs NLF-CHD5-AS, 74 colonies, 95% CI = 62 to 86 colonies, P < .001; IMR5-CHD5, 11 colonies, 95% CI = 2 to 20 colonies, vs IMR5-CHD5-AS, 39 colonies, 95% CI = 17 to 60 colonies, P = .01; tumor growth, n = 10 mice per group: mean tumor size at 5 weeks, NLF-CHD5, 0.36 cm3, 95% CI = 0.17 to 0.44 cm3, vs NLF-CHD5-AS, 1.65 cm3, 95% CI = 0.83 to 2.46 cm3, P = .002; IMR5-CHD5, 0.28 cm3, 95% CI = 0.18 to 0.38 cm3, vs IMR5-CHD5-AS, 1.15 cm3, 95% CI = 0.43 to 1.87 cm3; P = .01). High CHD5 expression was strongly associated with favorable event-free and overall survival (P < .001), even after correction for MYCN amplification and 1p deletion (P = .027).ConclusionsCHD5 is the strongest candidate tumor suppressor gene that is deleted from 1p36.31 in neuroblastomas, and inactivation of the second allele may occur by an epigenetic mechanism.