Constitutional mismatch repair-deficiency and whole-exome sequencing as the means of the rapid detection of the causative MSH6 defect.

BACKGROUND: Cases of children with more than one type of cancer either diagnosed simultaneously or successively, rarely occur in pediatric oncology. A second malignant neoplasm may be caused by mutagenic effects of the treatment of the primary malignancy and/or may point towards an underlying genetic cancer susceptibility syndrome. One example of such a syndrome is constitutional mismatch repair-deficiency, (CMMR-D) which carries an increased risk of various tumors including childhood hematologic malignancies and Lynch syndrome associated tumors. Timely diagnosis of CMMR-D is crucial, since this diagnosis has implications for the entire family. PATIENT: We report the case of a 15-year-old girl who was born to consanguineous parents. At the age of 20 months she was diagnosed with a T-cell non-Hodgkin lymphoma. Treatment was given according to NHL-BFM 95. 12 years later, an invasive adenocarcinoma of the colon was surgically removed which relapsed shortly afterwards. METHODS: Whole-exome sequencing of germline DNA was employed to rapidly detect the underlying mutation in this suspected CMMR-D patient. RESULTS: After a short turnaround time of less than 3 weeks, the diagnosis of CMMR-D could be confirmed by the identification of a homozygous 29-bp deletion in MSH6 (exon 6), which was confirmed by independent methods. CONCLUSIONS: We demonstrate that "bed-side" whole-exome sequencing is both feasible and cost-effective and may be the method of choice to rapidly uncover the genetical basis of (inherited) diseases.

The impact of TEL-AML1 (ETV6-RUNX1) expression in precursor B cells and implications for leukaemia using three different genome-wide screening methods.

The reciprocal translocation t(12;21)(p13;q22), the most common structural genomic alteration in B-cell precursor acute lymphoblastic leukaemia in children, results in a chimeric transcription factor TEL-AML1 (ETV6-RUNX1). We identified directly and indirectly regulated target genes utilizing an inducible TEL-AML1 system derived from the murine pro B-cell line BA/F3 and a monoclonal antibody directed against TEL-AML1. By integration of promoter binding identified with chromatin immunoprecipitation (ChIP)-on-chip, gene expression and protein output through microarray technology and stable labelling of amino acids in cell culture, we identified 217 directly and 118 indirectly regulated targets of the TEL-AML1 fusion protein. Directly, but not indirectly, regulated promoters were enriched in AML1-binding sites. The majority of promoter regions were specific for the fusion protein and not bound by native AML1 or TEL. Comparison with gene expression profiles from TEL-AML1-positive patients identified 56 concordantly misregulated genes with negative effects on proliferation and cellular transport mechanisms and positive effects on cellular migration, and stress responses including immunological responses. In summary, this work for the first time gives a comprehensive insight into how TEL-AML1 expression may directly and indirectly contribute to alter cells to become prone for leukemic transformation.

Loss-of-function mutations within the IL-2 inducible kinase ITK in patients with EBV-associated lymphoproliferative diseases.

The purpose of this study was the appraisal of the clinical and functional consequences of germline mutations within the gene for the IL-2 inducible T-cell kinase, ITK. Among patients with Epstein-Barr virus-driven lymphoproliferative disorders (EBV-LPD), negative for mutations in SH2D1A and XIAP (n=46), we identified two patients with R29H or D500T,F501L,M503X mutations, respectively. Human wild-type (wt) ITK, but none of the mutants, was able to rescue defective calcium flux in murine Itk(-/-) T cells. Pulse-chase experiments showed that ITK mutations lead to varying reductions of protein half-life from 25 to 69% as compared with wt ITK (107 min). The pleckstrin homology domain of wt ITK binds most prominently to phosphatidylinositol monophosphates (PI(3)P, PI(4)P, PI(5)P) and to lesser extend to its double or triple phosphorylated derivates (PIP2, PIP3), interactions which were dramatically reduced in the patient with the ITK(R29H) mutant. ITK mutations are distributed over the entire protein and include missense, nonsense and indel mutations, reminiscent of the situation in its sister kinase in B cells, Bruton's tyrosine kinase.

Concomitant treatment with urodilatin (ularitide) does not improve renal function in patients with acute renal failure after major abdominal surgery--a randomized controlled trial.

Acute renal failure after major abdominal surgery is a severe complication in critically ill patients in intensive care units (ICU). The aim of the study was to investigate the effect of urodilatin on the peak value and course of serum creatinine in patients with acute renal insufficiency after major abdominal surgery and the necessity of apparatus-based renal replacement treatment. Furthermore, the incidence and nature of adverse events under urodilatin was documented. In a prospective randomized double-blind placebo-controlled study, 12 critically ill patients after major abdominal surgery with acute renal failure in an intensive care unit (ICU) received 20 ng/kg b.w./min urodilatin (ularitide, INN) or placebo in addition to the standard diuretic therapy or low-dose dopamine (2.5 micrograms/kg b.w./min) and furosemide (1000 mg/24 hr) for a minimum of 96 hours. All patients received mechanical ventilation. Both groups had similar serum creatinine values on the day before the onset of urodilatin/placebo infusion (2.80 +/- 0.24 mg/dl, 2.93 +/- 0.48 mg/dl). Peak serum creatinine was lower in the urodilatin group (4.65 +/- 0.57 mg/dl) compared to vehicle treatment (5.78 +/- 1.58 mg/dl), although the difference did not reach statistical significance (P = 0.148). The total number of hemodialyses due to oligo-/anuria and/or hyperkalemia was the same in both groups during the study. In 4 patients of the placebo group, diuresis was reduced to anuria, whereas only 1 of the patients treated with urodilatin became anuric. No hemodynamic side effects or adverse events due to urodilatin were observed. This clinical study under double blind conditions revealed that the addition of urodilatin to the standard diuretic therapy of low-dose dopamine and furosemide failed to improve renal function in patients with established acute renal failure and that urodilatin did not eliminate the need for apparatus-based renal replacement treatment.