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Myelodysplastic/myeloproliferative neoplasms, unclassifiable (MDS/MPN-U) are rare genetically heterogeneous hematologic diseases associated with older age and a poor prognosis. If the disease progresses into acute myeloid leukemia (AML), it is often refractory to treatment. To gain insight into genetic alterations associated with disease progression, whole exome sequencing and single nucleotide polymorphism arrays were used to characterize the bone marrow and blood samples from a 39-year-old woman at MDS/MPN-U diagnosis and at AML progression, in which routine genetic diagnostics had not identified any genetic alterations. The data revealed the presence of a partial tandem duplication of the MLL gene as the only detectable copy number change and 11 non-silent somatic mutations, including DNMT3A R882H and NRAS G13D. All somatic lesions were present both at initial MDS/MPN-U diagnosis and at AML presentation at similar mutant allele frequencies. The patient has since had two extramedullary relapses and is at high risk of a future bone marrow relapse. A directed ex vivo drug sensitivity analysis showed that the patient's AML cells are sensitive to, for example, the MEK inhibitor trametinib and the proteasome inhibitor bortezomib, indicating that she may benefit from treatment with these drugs. © 2016 Wiley Periodicals, Inc.

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children and adolescents. Alveolar (ARMS) and embryonal (ERMS) histologies predominate, but rare cases are classified as spindle cell/sclerosing (SRMS). For treatment stratification, RMS is further subclassified as fusion-positive (FP-RMS) or fusion-negative (FN-RMS), depending on whether a gene fusion involving PAX3 or PAX7 is present or not. We investigated 19 cases of pediatric RMS using high resolution single-nucleotide polymorphism (SNP) array. FP-ARMS displayed, on average, more structural rearrangements than ERMS; the single FN-ARMS had a genomic profile similar to ERMS. Apart from previously known amplification (e.g., MYCN, CDK4, and MIR17HG) and deletion (e.g., NF1, CDKN2A, and CDKN2B) targets, amplification of ERBB2 and homozygous loss of ASCC3 or ODZ3 were seen. Combining SNP array with cytogenetic data revealed that most cases were polyploid, with at least one case having started as a near-haploid tumor. Further bioinformatic analysis of the SNP array data disclosed genetic heterogeneity, in the form of subclonal chromosomal imbalances, in five tumors. The outcome was worse for patients with FP-ARMS than ERMS or FN-ARMS (6/8 vs. 1/9 dead of disease), and the only children with ERMS showing intratumor diversity or with MYOD1 mutation-positive SRMS also died of disease. High resolution SNP array can be useful in evaluating genomic imbalances in pediatric RMS.

Transforming growth factor beta-1 (TGFB1)-induced gene expression was studied in five pancreatic carcinoma cell lines and one known TGFB1-sensitive cell line (HaCaT) by use of high-density filter-based cDNA microarrays representing over 4,000 human genes. The results indicate a complex cellular response to TGFB1 with 10% of the investigated genes showing altered expression after 3 or 48 hr of TGFB1 exposure. The tumor cell lines displayed a gradually inversed gene expression pattern, which correlated with reduced sensitivity to TGFB1, as compared to the HaCaT cell line. In the HaCaT cells, several proapoptotic genes showed increased expression in response to TGFB1, whereas the expression of antiapoptotic genes was decreased. In contrast, two pancreatic carcinoma cell lines, previously found to be growth stimulated by TGFB1, displayed an expression pattern opposite to that of these genes. Similarly, the expression of other functional groups of genes, such as cell cycle and transcription factor related genes, was almost completely reversed in these two tumor cell lines. Importantly, three of the five investigated pancreatic carcinoma cell lines responded to TGFB1, although they had SMAD4 inactivations, suggesting that the observed gene expression changes in these cell lines must be accomplished by SMAD-independent pathways.

Previous cytogenetic and comparative genomic hybridization (CGH) analyses have shown that the gain of chromosome arm 12p is frequent in pancreatic carcinomas. We investigated 15 pancreatic carcinoma cell lines using CGH, fluorescence in situ hybridization (FISH), and semiquantitative polymerase chain reaction (PCR) to characterize 12p amplifications in detail. The CGH analysis revealed gains of 12p in four of the cell lines and local amplification within 12p11-12 in six cell lines. By FISH analysis, using precisely mapped YAC clones, the commonly amplified region was found to be approximately 5 Mb. The amplified segment extended from YAC 753f12, covering the KRAS2 locus, to YAC 891f1, close to the centromere. A semiquantitative PCR methodology was used to estimate genomic copy numbers of 14 precisely mapped expressed sequence tags (ESTs) and sequence-tagged sites, located within this interval. The level of amplification ranged from two- to 12-fold. The produced gene copy profiles revealed a 3.5-Mb segment with various local amplifications. This region includes KRAS2 and ranges from D12S1617 to sts-N38796. Two of the cell lines (primary and metastatic tumor from the same patient) showed amplification peaks within the distal region of this segment, two had peaks within the proximal region, one showed subpeaks in both regions, and one displayed amplification of the entire region. Chromosome segment-specific cDNA array analysis of 29 expressed sequences within the whole interval between D12S1617 and sts-N38796 indicated overexpression of four ESTs, two corresponding to DEC2 and PPFIBP1, and two to ESTs with unknown function. Expression analysis of these and of KRAS2 showed specific overexpression in the six cell lines with local 12p amplifications. These findings indicate two target regions within the 3.5-Mb segment in 12p11-12, one proximal including PPFIBP1, and one distal including KRAS2.