Genome-wide profiling of follicular lymphoma by array comparative genomic hybridization reveals prognostically significant DNA copy number imbalances.

The secondary genetic events associated with follicular lymphoma (FL) progression are not well defined. We applied genome-wide BAC array comparative genomic hybridization to 106 diagnostic biopsies of FL to characterize regional genomic imbalances. Using an analytical approach that defined regions of copy number change as intersections between visual annotations and a Hidden Markov model-based algorithm, we identified 71 regional alterations that were recurrent in at least 10% of cases. These ranged in size from approximately 200 kb to 44 Mb, affecting chromosomes 1, 5, 6, 7, 8, 10, 12, 17, 18, 19, and 22. We also demonstrated by cluster analysis that 46.2% of the 106 cases could be sub-grouped based on the presence of +1q, +6p/6q-, +7, or +18. Survival analysis showed that 21 of the 71 regions correlated significantly with inferior overall survival (OS). Of these 21 regions, 16 were independent predictors of OS using a multivariate Cox model that included the international prognostic index (IPI) score. Two of these 16 regions (1p36.22-p36.33 and 6q21-q24.3) were also predictors of transformation risk and independent of IPI. These prognostic features may be useful to identify high-risk patients as candidates for risk-adapted therapies.

High resolution analysis of follicular lymphoma genomes reveals somatic recurrent sites of copy-neutral loss of heterozygosity and copy number alterations that target single genes.

A multiplatform approach, including conventional cytogenetic techniques, BAC array comparative genomic hybridization, and Affymetrix 500K SNP arrays, was applied to the study of the tumor genomes of 25 follicular lymphoma biopsy samples with paired normal DNA samples to characterize balanced translocations, copy number imbalances, and copy-neutral loss of heterozygosity (cnLOH). In addition to the t(14;18), eight unique balanced translocations were found. Commonly reported FL-associated copy number regions were revealed including losses of 1p32-36, 6q, and 10q, and gains of 1q, 6p, 7, 12, 18, and X. The most frequent regions affected by copy-neutral loss of heterozygosity were 1p36.33 (28%), 6p21.3 (20%), 12q21.2-q24.33 (16%), and 16p13.3 (24%). We also identified by SNP analysis, 45 aberrant regions that each affected one gene, including CDKN2A, CDKN2B, FHIT, KIT, PEX14, and PTPRD, which were associated with canonical pathways involved in tumor development. This study illustrates the power of using complementary high-resolution platforms on paired tumor/normal specimens and computational analysis to provide potential insights into the significance of single-gene somatic aberrations in FL tumorigenesis.

The significance of TP53 in lymphoid malignancies: mutation prevalence, regulation, prognostic impact and potential as a therapeutic target.

The tumour suppressor TP53 (previously termed p53) mediates a pathway that is considered to be one of the most important mechanisms in the maintenance of genomic stability. The function of TP53 can be abrogated by genomic deletion, mutation, or deregulation of upstream and downstream participants in the TP53 pathway. While aberrations of TP53 are widely prevalent in non-haematological malignancies (over 60%), they are present in much lower frequency in haematological malignancies (<20%). Nevertheless, in those cases where TP53 function or expression is aberrant, correlation with inferior clinical outcome (such as overall survival and progression or transformation) has generally been strong. In this review, we focus our discussion on the relationship between TP53 and lymphoid malignancies as defined by the World Health Organization. Specifically, we examine the prevalence of TP53 aberrations and their prognostic significance in various types of lymphoid cancer. Next, we discuss the various mechanisms of TP53 inactivation. Finally, we summarize progress in the use of recent therapeutic modalities that target TP53.

Tissue-specific regulation of Fas/APO-1/CD95 expression by p53.

The regulation of Fas/APO-1(CD95), an important member of the tumor necrosis factor (TNF) superfamily involved in membrane-mediated apoptosis, has been a subject of recent research. Ligation of Fas by Fas ligand or an anti-Fas cross-linking antibody triggers receptor trimerization followed by recruitment of FADD to the cytoplasmic domain of the receptor and the activation of the caspase cascade. The tumor suppressor p53 has been shown to upregulate Fas expression under numerous pro-apoptotic stimuli in vitro. Using the p53 knockout mouse model, we demonstrate by Western blot analysis, immunohistochemistry, and semi-quantitative RT-PCR that Fas expression is reduced in spleen and liver from p53-/- mice compared to p53+/+ controls, while similar expression levels were observed in brain, heart, kidney, lung, skin, testis, and thymus between the two groups. While Fas protein was abundant in brain, heart, liver, and spleen, low levels of endogenous expression was observed in other tissues from the p53+/+ and p53-/- mice. These data indicate that p53 regulates Fas expression in a tissue-specific manner.

Utility of array comparative genomic hybridization in cytogenetic analysis.

Conventional comparative genomic hybridization (CGH), high-resolution oligonucleotide, and BAC array CGH have modernized the field of cytogenetics to enable access to unbalanced genomic aberrations such as whole or partial chromosomal gains and losses. The basic principle of array CGH involves hybridizing differentially labeled proband/test (e.g., tumor) and normal reference DNA on an array of oligonucleotide or BAC clones instead of normal metaphases as in conventional CGH. The sub-megabase resolution tiling BAC arrays are extremely useful for the analysis of acquired aberrations in cancer genomes. Array CGH can be extremely useful to identify the chromosomal makeup of marker and ring chromosomes, to define/delineate the precise location/bands involved in structural aberrations and the accurate localization of translocation breakpoints in both simple and complex karyotypes either alone or in combination with standard karyotype analysis.

Acquired TNFRSF14 mutations in follicular lymphoma are associated with worse prognosis.

Clinical correlative studies have linked 1p36 deletions with worse prognosis in follicular lymphoma (FL). In this study, we sought to identify the critical gene(s) in this region that is responsible for conferring inferior prognosis. BAC array technology applied to 141 FL specimens detected a minimum region of deletion (MRD) of ∼97 kb within 1p36.32 in 20% of these cases. Frequent single-nucleotide polymorphism-detected copy-neutral loss of heterozygosity was also found in this region. Analysis of promoter CpGs in the MRD did not reveal differential patterns of DNA methylation in samples that differed in 1p36 status. Exon sequencing of MRD genes identified somatic alterations in the TNFRSF14 gene in 3 of 11 selected cases with matching normal DNA. An expanded cohort consisting of 251 specimens identified 46 cases (18.3%) with nonsynonymous mutations affecting TNFRSF14. Overall survival (OS) and disease-specific survival (DSS) were associated with the presence of TNFRSF14 mutation in patients whose overall treatment included rituximab. We further showed that inferior OS and DSS were most pronounced in patients whose lymphomas contained both TNFRSF14 mutations and 1p36 deletions after adjustment for the International Prognostic Index [hazard ratios of 3.65 (95% confidence interval, 1.35-9.878, P=0.011) and 3.19 (95% confidence interval, 1.06-9.57, P=0.039), respectively]. Our findings identify TNFRSF14 as a candidate gene associated with a subset of FL, based on frequent occurrence of acquired mutations and their correlation with inferior clinical outcomes.

Tyrosine hydroxylase expression and Cdk5 kinase activity in ataxic cerebellum.

Ataxia has been associated with abnormalities in neuronal differentiation and migration, which are regulated by Cyclin-dependent kinase 5 (Cdk5). The cerebellum of mice lacking Cdk5 or its activator, p35, resembles those of ataxic reeler and scrambler mice, suggesting that Cdk5 may contribute to ataxic pathology. As with other ataxic mice, the pogo/pogo mouse shows aberrant cerebellar tyrosine hydroxylase (TH) expression. Since Cdk5 phosphorylates and upregulates TH expression, we sought to analyze (i) Cdk5 activity in the pogo cerebellum, which exhibits abnormal TH expression, and (ii) TH expression in the cerebellum of p35-/- and p39-/- mice, which display reduced Cdk5 activity. Interestingly, we found that increased TH expression in the pogo cerebellum coincided with reduced Cdk5 activity. However, reduced Cdk5 activity in both p35-/- and p39-/- cerebellum did not correspond to defects in TH expression. Together, these suggest that abnormal TH expression in the cerebellum might be regulated by mechanisms other than Cdk5 activity.