Paradoxical down-regulation of p16INK4a mRNA with advancing age in Acute Myeloid Leukemia

Aging is generally considered to be the consequence of stem cell attrition caused by the activity of tumor suppressor pathways that censor potentially malignant clones by eliciting apoptosis or senescence. An important effector of aging is the cyclindependent kinase inhibitor p16INK4a, which is also a known suppressor of cancer. The expression of p16INK4a is very low or absent in young organisms but increases with advancing age. We recently showed that, unlike healthy cells, acute myeloid leukemia (AML) derived blasts show a down-regulation of p16INK4a mRNA with increasing age. Based on this observation we hypothesize that suppression of defense mechanisms which protect older cells against cellular and DNA damage might facilitate oncogenesis in older individuals

A single center analysis of nucleophosmin in acute myeloid leukemia:value of combining immunohistochemistry with molecular mutation analysis

Mutations of nucleophosmin 1 are frequently found in acute myeloid leukemia and lead to aberrant cytoplasmic accumulation of nucleophosmin protein. Immunohistochemical staining is therefore recommended as the technique of choice in front-line screening. In this study, we assessed the sensitivity and specificity of immunohistochemistry on formalin-fixed bone marrow biopsies compared with gold standard molecular analysis to predict nucleophosmin 1 mutation status in 119 patients with acute myeloid leukemia. Discrepant cases were further characterized by gene expression analyses and fluorescence in situ hybridization. A large overlap between both methods was observed. Nevertheless, nine patients demonstrated discordant results at initial screening. Five cases demonstrated nuclear staining of nucleophosmin 1 by immunohistochemistry, but a nucleophosmin 1 mutation by molecular analysis. In two cases this could be attributed to technical issues and in three cases minor subpopulations of myeloblasts had not been discovered initially. All tested cases exhibited the characteristic nucleophosmin-mutated gene expression pattern. Four cases had cytoplasmic nucleophosmin 1 staining and a nucleophosmin-mutated gene expression pattern without a detectable nucleophosmin 1 mutation. In two of these cases we found the chromosomal translocation t(3;5)(q25;q35) encoding the NPM-MLF1 fusion protein. In the other discrepant cases the aberrant cytoplasmic nucleophosmin staining and gene expression could not be explained. In total six patients (5%) had true discordant results between immunohistochemistry and mutation analysis. We conclude that cytoplasmic nucleophosmin localization is not always caused by a conventional nucleophosmin 1 mutation and that in the screening for nucleophosmin 1 abnormalities, most information will be obtained by combining immunohistochemistry with molecular analysis

CombiFlow:combinatorial AML-specific plasma membrane expression profiles allow longitudinal tracking of clones

Acute myeloid leukemia (AML) often presents as an oligoclonal disease whereby multiple genetically distinct subclones can coexist within patients. Differences in signaling and drug sensitivity of such subclones complicate treatment and warrant tools to identify them and track disease progression. We previously identified >50 AML-specific plasma membrane (PM) proteins, and 7 of these (CD82, CD97, FLT3, IL1RAP, TIM3, CD25, and CD123) were implemented in routine diagnostics in patients with AML (n = 256) and myelodysplastic syndrome (n = 33). We developed a pipeline termed CombiFlow in which expression data of multiple PM markers is merged, allowing a principal component–based analysis to identify distinctive marker expression profiles and to generate single-cell t-distributed stochastic neighbor embedding landscapes to longitudinally track clonal evolution. Positivity for one or more of the markers after 2 courses of intensive chemotherapy predicted a shorter relapse-free survival, supporting a role for these markers in measurable residual disease (MRD) detection. CombiFlow also allowed the tracking of clonal evolution in paired diagnosis and relapse samples. Extending the panel to 36 AML-specific markers further refined the CombiFlow pipeline. In conclusion, CombiFlow provides a valuable tool in the diagnosis, MRD detection, clonal tracking, and understanding of clonal heterogeneity in AML

Aging of hematopoietic stem cells:Intrinsic changes or micro-environmental effects?

During development hematopoietic stem cells (HSCs) expand in number and persist throughout life by undergoing self-renewing divisions. Nevertheless, the hematopoietic system does not escape the negative effects of aging, suggesting that self-renewal is not complete. A fundamental issue in stem cell biology relates to such age-dependent loss of stem cell activity. Both stem cell intrinsic factors and extrinsic factors associated with an aging micro-environment could contribute to aging of the hematopoietic system. Recently, changes in the clonal composition of the HSC compartment during aging have been put forward as a key factor. Here, we discuss these recent developments and speculate how they may be of clinical relevance

Loss of quiescence and impaired function of CD34(+)/CD38(low) cells one year following autologous stem cell transplantation

<p>Patients who have undergone autologous stem cell transplantation are subsequently more susceptible to chemotherapy-induced bone marrow toxicity. In the present study, bone marrow primitive progenitor cells were examined one year after autologous stem cell transplantation and compared with normal bone marrow and mobilized peripheral blood stem cells. Post-transplantation bone marrow contained a significantly lower percentage of quiescent cells in the CD34(+)/CD38(low) fraction compared to normal bone marrow. In addition, we observed a strong decrease in stem cell/primitive progenitor frequency in post-transplantation CD34(+) cells as defined by long-term culture assays. Measurement of the levels of reactive oxygen species by flow cytometry revealed comparable levels in post-transplantation and normal bone marrow CD34(+)/CD38(low) cells, while significantly higher levels of reactive oxygen species were observed in CD34(+)/CD38(high) cells following autologous stem cell transplantation compared to normal bone marrow. Moreover, post-transplantation CD34(+) bone marrow cells demonstrated an increased sensitivity to buthionine sulfoximine, a trigger for endogenous production of reactive oxygen species. Gene expression analysis on CD34(+) cells revealed a set of 195 genes, including HMOX1, EGR1, FOS and SIRPA that are persistently down-regulated in mobilized peripheral blood cells and post-transplantation bone marrow compared to normal bone marrow. In conclusion, our data indicate that the diminished regenerative capacity of bone marrow following autologous stem cell transplantation is possibly related to a loss of quiescence and a reduced tolerability to oxidative stress.</p>