ARACNE: An Algorithm for the Reconstruction of Gene Regulatory Networks in a Mammalian Cellular Context

Elucidating gene regulatory networks is crucial for understanding normal cell physiology and complex pathologic phenotypes. Existing computational methods for the genome-wide "reverse engineering" of such networks have been successful only for lower eukaryotes with simple genomes. Here we present ARACNE, a novel algorithm, using microarray expression profiles, specifically designed to scale up to the complexity of regulatory networks in mammalian cells, yet general enough to address a wider range of network deconvolution problems. This method uses an information theoretic approach to eliminate the majority of indirect interactions inferred by co-expression methods. We prove that ARACNE reconstructs the network exactly (asymptotically) if the effect of loops in the network topology is negligible, and we show that the algorithm works well in practice, even in the presence of numerous loops and complex topologies. We assess ARACNE's ability to reconstruct transcriptional regulatory networks using both a realistic synthetic dataset and a microarray dataset from human B cells. On synthetic datasets ARACNE achieves very low error rates and outperforms established methods, such as Relevance Networks and Bayesian Networks. Application to the deconvolution of genetic networks in human B cells demonstrates ARACNE's ability to infer validated transcriptional targets of the cMYC proto-oncogene. We also study the effects of misestimation of mutual information on network reconstruction, and show that algorithms based on mutual information ranking are more resilient to estimation errors. ARACNE shows promise in identifying direct transcriptional interactions in mammalian cellular networks, a problem that has challenged existing reverse engineering algorithms. This approach should enhance our ability to use microarray data to elucidate functional mechanisms that underlie cellular processes and to identify molecular targets of pharmacological compounds in mammalian cellular networks

MicroRNA expression in HTLV-1 infection and pathogenesis

Our laboratory is examining the profiles of microRNA expression in ATLL cells and infected T-cell lines using microarrays and small RNA libraries. Microarray analysis of ATLL samples revealed 6 upregulated and 21 downregulated microRNAs in ATLL cells compared to CD4+ T-cell controls. Potential targets for deregulated microRNAs were identified by integrating microRNA and mRNA expression profiles. Current experiments are aimed at verifying these predicted microRNA-target interactions

Identification of rare Epstein-Barr virus infected memory B cells and plasma cells in non-monomorphic post-transplant lymphoproliferative disorders and the signature of viral signaling

Background and Objectives. In early and polymorphic post-transplant lymphoprolifera- tive disorders (PTLD) Epstein-Barr virus (EBV), through its latency proteins, drives the proliferation of B lymphocytes, a process which in immunocompetent individuals leads to the establishment of latently infected memory B cells. Design and Methods. We analyzed 11 cases, which included early and polymorphic PTLD, and 12 controls for latency of EBV infection and their antigenic profile. Results. We identified a minority of terminally differentiated EBER+ IRTA1+ memory B cells and EBER+ CD138+ PRDM1+ plasma cells in these samples. These elements were identified both in PTLD and in tumor-free tonsils from post-transplant patients but not in EBV– control tonsils. The expression of EBV latency proteins is heterogeneous, and is associated with activation of the NF-κB pathway. EBV signaling (through EBNA2, LMP1 and LMP2A) and NF-κB activation correlated with upregulation of target proteins: cMYC, JunB, CCL22, TRAF1 and IRF4. EBV-infected lymphocytes in early and polymor- phic PTLDs represent a mixture of latencies II, III and, in at least 1/3 of infected cells, of latency 0. Interpretation and conclusions. EBV infection correlates with NF-κB activation, with EBV-dependent cell signaling, and lastly, with the presence of EBV-infected plasma cells and memory cells. Key words: post-transplant lymphoproliferative disorder, Epstein-Barr virus, viral latency, NF-κB signaling, plasma cell, memory B cell

Retrospective Study of Adverse Events of Chemotherapy in Cats

Background: Clients who seek veterinary care for pets with cancer are often concerned about the potential negative impact of chemotherapeutic treatments on their animals’ quality of life. A consensus currently exists in veterinary oncology regarding the quantification and rating of adverse treatment effects in dogs and cats in response to chemotherapy agents. This grading system is referred to as Veterinary Cooperative Oncology Group - Common Terminology Criteria for Adverse Events. The purpose of this retrospective case series was to investigate the delayed acute effects of chemotherapy drugs in cats receiving cancer treatment.Materials, Methods & Results: Medical records were reviewed to determine the chemotherapy agent used and delayed adverse effects. Side effects were classified according to Veterinary Co-operative Oncology Group grading. All cats were evaluated after the first chemotherapy administration, after a single dose. The reported effects included hematologic effects (e.g., neutropenia, thrombocytopenia, increases in liver enzymes, and azotemia), gastrointestinal effects (e.g., vomiting, diarrhea, and inappetence), and sepsis. All of the cats in this study received ondansetron and omeprazol  in the first five days following chemotherapy administration. If vomiting occurred with oral medication, maropitant was administered subcutaneously for three consecutive days. If diarrhea (> grade II) occurred, probiotics were administered for seven days. Hematologic examination was performed 3-14 days after chemotherapy. If neutropenia (> grade III) occurred, Human granulocyte colony stimulating factor was administered subcutaneously for three consecutive days together with prophylactic antibiotics. Lomustine, carboplatin, vincristine, doxorubicin, cyclophosphamide, mitoxantrone, and vinblastine were administered in 33%, 19%, 16%, 5%, 16%, 10% and 2% of the cases examined, respectively. The most common adverse events were vomiting, inappetence, neutropenia, and thrombocytopenia. Vomiting occurred in 6% cases, most of them associated with cyclophosphamide. Inappetence/anorexia affected 12% of the cases, mostly those involving cyclophosphamide or doxorubicin. Neutropenia was observed in 22% of the cases, with cyclophosphamide, followed by carboplatin and lomustine. According to the current grading system of adverse effects induced by chemotherapy, grade I toxicity was observed in 83% of the cases, whiles grade II-IV were observed in 7%, 8%, and 2% of the cases examined, respectively.Discussion: In general, the chemotherapy regimens in the cases examined were well tolerated. The toxicity experienced was infrequent and mostly mild, thereby resulting in satisfactory tolerability of the chemotherapy regimens. According to the current grading system for the adverse effects of chemotherapy, 83% of the cases examined included grade I, indicating that most of the cats experienced asymptomatic, or mild symptoms, and medical intervention was not needed. In previous studies of dogs and cats, a severe adverse event following chemotherapy was reported for fewer than 1 in 4 animals, and approximately 3-5% experienced a serious adverse event that led to hospitalization. In the present study, 2% of the cats experienced serious or life threatening adverse events. The only chemotherapeutic agent that was associated with inappetence, vomiting, and neutropenia was cyclophosphamide. Based on the data examined, we would recommend that cyclophosphamide should be used with caution for the treatment of cancer in cats, with adequate antiemetic and nutritional support available if needed. In addition, febrile neutropenia/sepsis may be avoided by using a fractionated schedule

A systems biology approach to prediction of oncogenes and molecular perturbation targets in B-cell lymphomas

The computational identification of oncogenic lesions is still a key open problem in cancer biology. Although several methods have been proposed, they fail to model how such events are mediated by the network of molecular interactions in the cell. In this paper, we introduce a systems biology approach, based on the analysis of molecular interactions that become dysregulated in specific tumor phenotypes. Such a strategy provides important insights into tumorigenesis, effectively extending and complementing existing methods. Furthermore, we show that the same approach is highly effective in identifying the targets of molecular perturbations in a human cellular context, a task virtually unaddressed by existing computational methods. To identify interactions that are dysregulated in three distinct non-Hodgkin's lymphomas and in samples perturbed with CD40 ligand, we use the B-cell interactome (BCI), a genome-wide compendium of human B-cell molecular interactions, in combination with a large set of microarray expression profiles. The method consistently ranked the known gene in the top 20 (0.3%), outperforming conventional approaches in 3 of 4 cases

Gene Expression Profiling of Hairy Cell Leukemia Reveals a Phenotype Related to Memory B Cells with Altered Expression of Chemokine and Adhesion Receptors

Hairy cell leukemia (HCL) is a chronic B cell malignancy characterized by the diffuse infiltration of bone marrow and spleen by cells displaying a typical “hairy” morphology. However, the nature of the HCL phenotype and its relationship to normal B cells and to other lymphoma subtypes remains unclear. Using gene expression profiling, we show here that HCL displays a homogeneous pattern of gene expression, which is clearly distinct from that of other B cell non-Hodgkin lymphomas. Comparison with the gene expression profiles of purified normal B cell subpopulations, including germinal center (GC), pre-GC (naive), and post-GC (memory) B cells, shows that HCL cells are more related to memory cells, suggesting a derivation from this B cell population. Notably, when compared with memory cells, HCL cells displayed a remarkable conservation in proliferation, apoptosis, and DNA metabolism programs, whereas they appeared significantly altered in the expression of genes controlling cell adhesion and response to chemokines. Finally, these analyses have identified several genes that are specifically expressed in HCL and whose expression was confirmed at the protein level by immunocytochemical analysis of primary HCL cases. These results have biological implications relevant to the pathogenesis of this malignancy as well as clinical implications for its diagnosis and therapy