Combined mutations of ASXL1, CBL, FLT3, IDH1, IDH2, JAK2, KRAS, NPM1, NRAS, RUNX1, TET2 and WT1 genes in myelodysplastic syndromes and acute myeloid leukemias

<p>Abstract</p> <p>Background</p> <p>Gene mutation is an important mechanism of myeloid leukemogenesis. However, the number and combination of gene mutated in myeloid malignancies is still a matter of investigation.</p> <p>Methods</p> <p>We searched for mutations in the <it>ASXL1, CBL, FLT3, IDH1, IDH2, JAK2, KRAS, NPM1, NRAS, RUNX1, TET2 </it>and <it>WT1 </it>genes in 65 myelodysplastic syndromes (MDSs) and 64 acute myeloid leukemias (AMLs) without balanced translocation or complex karyotype.</p> <p>Results</p> <p>Mutations in <it>ASXL1 </it>and <it>CBL </it>were frequent in refractory anemia with excess of blasts. Mutations in <it>TET2 </it>occurred with similar frequency in MDSs and AMLs and associated equally with either <it>ASXL1 </it>or <it>NPM1 </it>mutations. Mutations of <it>RUNX1 </it>were mutually exclusive with <it>TET2 </it>and combined with <it>ASXL1 </it>but not with <it>NPM1</it>. Mutations in <it>FLT3 (</it>mutation and internal tandem duplication), <it>IDH1</it>, <it>IDH2</it>, <it>NPM1 </it>and <it>WT1 </it>occurred primarily in AMLs.</p> <p>Conclusion</p> <p>Only 14% MDSs but half AMLs had at least two mutations in the genes studied. Based on the observed combinations and exclusions we classified the 12 genes into four classes and propose a highly speculative model that at least a mutation in one of each class is necessary for developing AML with simple or normal karyotype.</p

Prediction of Co-Receptor Usage of HIV-1 from Genotype

Human Immunodeficiency Virus 1 uses for entry into host cells a receptor (CD4) and one of two co-receptors (CCR5 or CXCR4). Recently, a new class of antiretroviral drugs has entered clinical practice that specifically bind to the co-receptor CCR5, and thus inhibit virus entry. Accurate prediction of the co-receptor used by the virus in the patient is important as it allows for personalized selection of effective drugs and prognosis of disease progression. We have investigated whether it is possible to predict co-receptor usage accurately by analyzing the amino acid sequence of the main determinant of co-receptor usage, i.e., the third variable loop V3 of the gp120 protein. We developed a two-level machine learning approach that in the first level considers two different properties important for protein-protein binding derived from structural models of V3 and V3 sequences. The second level combines the two predictions of the first level. The two-level method predicts usage of CXCR4 co-receptor for new V3 sequences within seconds, with an area under the ROC curve of 0.937±0.004. Moreover, it is relatively robust against insertions and deletions, which frequently occur in V3. The approach could help clinicians to find optimal personalized treatments, and it offers new insights into the molecular basis of co-receptor usage. For instance, it quantifies the importance for co-receptor usage of a pocket that probably is responsible for binding sulfated tyrosine

Frequent CXCR4 tropism of HIV-1 subtype A and CRF02_AG during late-stage disease - indication of an evolving epidemic in West Africa

<p>Abstract</p> <p>Background</p> <p>HIV-1 is one of the fastest evolving pathogens, and is distinguished by geographic and genetic variants that have been classified into different subtypes and circulating recombinant forms (CRFs). Early in infection the primary coreceptor is CCR5, but during disease course CXCR4-using HIV-1 populations may emerge. This has been correlated with accelerated disease progression in HIV-1 subtype B. Basic knowledge of HIV-1 coreceptor tropism is important due to the recent introduction of coreceptor antagonists in antiretroviral therapy, and subtype-specific differences regarding how frequently HIV-1 CXCR4-using populations appear in late-stage disease need to be further investigated. To study how frequently CXCR4-using populations appear in late-stage disease among HIV-1 subtype A and CRF02_AG, we evaluated the accuracy of a recombinant virus phenotypic assay for these subtypes, and used it to determine the HIV-1 coreceptor tropism of plasma samples collected during late-stage disease in Guinea-Bissau. We also performed a genotypic analysis and investigated subtype-specific differences in the appearance of CXCR4 tropism late in disease.</p> <p>Results</p> <p>We found that the recombinant virus phenotypic assay accurately predicted HIV-1 coreceptor tropism of subtype A and CRF02_AG. Over the study period (1997-2007), we found an increasing and generally high frequency of CXCR4 tropism (86%) in CRF02_AG. By sequence analysis of the V3 region of our samples we developed a novel genotypic rule for predicting CXCR4 tropism in CRF02_AG, based on the combined criteria of the total number of charged amino acids and net charge. This rule had higher sensitivity than previously described genotypic rules and may be useful for development of future genotypic tools for this CRF. Finally, we conducted a literature analysis, combining data of 498 individuals in late-stage disease, and found high amounts of CXCR4 tropism for all major HIV-1 subtypes (60-77%), except for subtype C (15%).</p> <p>Conclusions</p> <p>The increase in CXCR4 tropism over time suggests an evolving epidemic of CRF02_AG. The results of the literature analysis demonstrate the need for further studies investigating subtype-specific emergence for CXCR4-tropism; this may be particularly important due to the introduction of CCR5-antagonists in HIV treatment regimens.</p

Bone marrow-derived macrophage production

Macrophages are critical components of the innate and adaptive immune responses, and they are the first line of defense against foreign invaders because of their powerful microbicidal activities. Macrophages are widely distributed throughout the body and are present in the lymphoid organs, liver, lungs, gastrointestinal tract, central nervous system, bone, and skin. Because of their repartition, they participate in a wide range of physiological and pathological processes. Macrophages are highly versatile cells that are able to recognize microenvironmental alterations and to maintain tissue homeostasis. Numerous pathogens have evolved mechanisms to use macrophages as Trojan horses to survive, replicate in, and infect both humans and animals and to propagate throughout the body. The recent explosion of interest in evolutionary, genetic, and biochemical aspects of host-pathogen interactions has renewed scientific attention regarding macrophages. Here, we describe a procedure to isolate and cultivate macrophages from murine bone marrow that will provide large numbers of macrophages for studying host-pathogen interactions as well as other processes

Effects of Coxiella burnetii on MAPKinases phosphorylation

Q fever is a disease caused by Coxiella burnetii, an obligate intracellular bacterium. Acute Q fever is characterized by efficient immune response, whereas chronic Q fever is characterized by dysregulated immune response as demonstrated by the lack of granulomas, the failure of C. burnetii to induce lymphoproliferation, and interferon-γ production. The mitogen-activated protein kinase (MAPK) signaling pathway plays crucial roles in innate immune responses and control of bacterial infections. However, its role in Q fever has not been addressed. First, we investigated the activation of MAPKs p38, c-jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) 1/2 in murine macrophages stimulated with C. burnetii. Coxiella burnetii NM phase I (virulent) and NM phase II (avirulent) induced the activation of JNK and ERK1/2. Avirulent C. burnetii activate p38, whereas C. burnetii did not induce the phosphorylation of p38. Second, the level of p38 activation was studied in Q fever patients. We found that p38 was activated in monocyte-derived macrophages from healthy donors and patients with acute Q fever in response to a potent agonist such as lipopolysaccharide. Interestingly, p38 was not activated in patients with active chronic Q fever and was activated in patients with cured chronic Q fever. These results suggest that the determination of p38 activation may serve as a tool for measuring Q fever activity