Class of 2013 Abstract / Specific Aims: The purpose of this study was to assess risk factors that are associated with inpatient charges and mortality rates with methicilin-resistant Staphylococcus aureu (MRSA), Aspergillus, and Candida in children with leukemia. The hypotheses is MRSA and opportunistic fungal infections are associated with higher inpatient charges and worse outcomes relative to those without. Children with leukemia are especially at risk due to underdeveloped and compromised immune systems. The rational is that identifying the risk factors that affect mortality and inpatient charges of these infections should add to the current knowledge of treating and preventing these infections in immune compromised patients. Methods: Retrospective cohort study using the Agency for Healthcare Research Quality (AHRQ) KIDS 2009 database. Inclusion criteria was defined as 17 years of age or younger and a ICD-9 code for an active infection with MRSA, Aspergillus, or Candida. Regression analysis's were performed to identify factors that had an impact on mortality, length of stay, and direct patient charges. Main Results: Values that were significant (p≤0.05) for predicting an increase in mortality were Age (year), MRSA, Aspergillus, Candida, and Deyo-Charlson comorbidity scores. Significant predictors of increased Inpatient Charges (p≤0.05) was Age (year), MRSA, Aspergillus, Candida, Female Sex, Deyo-Charlson comorbidity scores, Urban Hospitals, and the Southern Region. Significant predictors of Increased Length of Stay (p≤0.05) MRSA, Aspergillus, Candida, Female Sex, Deyo-Charlson, Urban Hospitals, Teaching Hospitals, and the Southern Region. Conclusion: Although uncommon in leukemia cases involving pediatrics or young adults, statistically significant and large risks of higher mortality, length of stay, and inpatient charges were noted in cases involving MRSA, aspergillus, and candida. Increasing Deyo-Charlson comorbidities scores were also consistently important predictors for poor outcomes in these leukemia patients and, with certain outcomes, increasing age and female sex.

Class of 2012 Abstract / Specific Aims: The objectives of this study were to determine the financial impact and mortality of CLL and Richter’s transformation in CLL in the inpatient setting in the payer’s perspective, the common diagnoses at discharge for patients with CLL, and to compare demographics, hospital characteristics, and co-morbidities for CLL cases versus Richter’s only cases. Methods: This study was a retrospective cohort of inpatient hospital charges and mortality of CLL patients and CLL patients with Richter’s transformation in the United States in the perspective of the payer. Using weighted statistical methods, results of this investigation yielded nationally-representative findings. The hospital charges were analyzed with a gamma regression with log link, and mortality was analyzed with a generalized linear regression. Main Results: There were total of 391,287 cases and 7% (27,259) were Richter’s cases. The overall hospital charges for CLL and CLL patients with Richter’s transformation from 2005 to 2009 were $38,735 (±58859) per case and $53,118 (±77993) per case, respectively. The mortality was 6.3% (24,520 deaths) overall and 9.1% mortality (2,485 deaths) for Richter’s transformation patients. The significant predictors (p < 0.05) that were associated with an increase the hospital charges for Richter’s patients was sepsis while sepsis and weight loss were associated with an increase in mortality. Conclusions This study adds to the few studies published to show the impact of CLL and Richter’s. However, due to the limitation on pharmacotherapies, it was not possible to determine therapeutic cost drivers for these cases. Future studies are warranted to determine the cost of therapies associated to the different stages of CLL.

No description available.

This thesis describes an attempt to increase the life span of mice after injection of a leukemic tumor. Fatty acids were used as a protecting agent against the leukemic tumor.

The oncogene Ahi-1 was recently identified through provirus insertional mutagenesis in murine leukemias and lymphomas. Its involvement in human leukemogenesis is demonstrated by gross perturbations in its expression in several leukemic cells lines, particularly in cutaneous T-cell lymphoma (CTCL) cell lines (Hut 78 and Hut 102). Hut 78 is derived from a patient with Sezary syndrome, a common leukemic variant of the human CTCL mycosis fungoides. Aberrant expression of AHI-1 mRNA and protein has been found in CD4⁺CD7⁻ leukemic Sezary cells from patients with Sezary syndrome. Moreover, stable suppression of AHI-1 using retroviral-mediated RNA interference in Hut 78 cells inhibits their transforming activity in vitro and in vivo. In an effort to identify genes involved in AHI-1-mediated leukemic transformation in CTCL, microarray analysis was performed to compare six RNA samples from AHI-1 suppressed Hut 78/sh4 cells to five samples from Hut 78 control cells. Limma and dChip analyses identified 218 and 95 differentially expressed genes, respectively, using a fold change criteria of > or < 2 and a p-value threshold of ≤ 0.01. After evaluation of both analyses, 21 genes were selected based upon interesting structural and functional information, specificity to hematopoietic cells or T-cells, and previous connections to cancer. Expression patterns of these 21 genes were validated by qRT-PCR with p-values < 0.05 ranging from 1.97 x 10⁻¹⁰ to 6.55 x 10⁻³, with the exception of BRDG1 at 5.88 x 10⁻². The observed up-regulation of both BIN1 and HCK in AHI-1 suppressed Hut 78/sh4 cells as compared to control cells further confirmed at the protein level. The tumor suppressor BIN1 is known to physically interact with c-MYC, which also exhibits differential protein expression in these cells. Characterization of BIN1 identified 4 isoforms all of which contain exon 10 and demonstrate alternative splicing of exons 12A and 13. Additionally, qRT-PCR results from primary Sezary samples indicate there is clinical significance in the expression changes detected for BIN1, HCK, REPS2, BRDG1, NKG7 and SPIB. These findings identify several new differentially expressed genes that may play critical roles in AHI-1-mediated leukemic transformation of human CTCL cells. / Medicine, Faculty of / Medical Genetics, Department of / Graduate

Outcomes for most patients with acute leukemia remain dismal. In-vitro, vesiculovirus members induced rapid apoptosis of acute leukemia cells. Intravenous injection of lymphoblastic leukemia cells infected ex-vivo with attenuated Vesicular Stomatitis Virus or Maraba Virus followed by gamma-irradiation, controlled leukemic progression in murine recipients. Essential properties of this autologous vaccine [immunotherapy by Leukemia-Oncotropic Virus (iLOV)] and the host’s immune system were characterized. iLOV durability was restricted to the leukemia used to manufacture the vaccine. At administration, virion cell-entry was required but vesiculovirus lifecycle completion was not essential. Apoptotic or necrotic leukemia cells, with/without co-injection of virus, were ineffective vaccines. Similarly ineffective were leukemia cells activated by, or injected with, Toll-like receptor agonists. Naïve recipients of adoptive splenocyte transfer from vaccine-treated immunocompetent donors were protected from leukemic challenge. Efficacy was notably diminished following matched allogeneic bone marrow transplantation; this correlated with isolated depletion of cytotoxic T-cells. iLOV was ineffective in athymic mice. Taken together, iLOV therapy relies on immediate spaciotemporal interactions between infected-dead/dying leukemia cells and the immune system; this promotes adaptive anti-tumor responses. Clinical translation could target patients in remission to control relapse. During the above I discovered that under specific conditions, live vesiculovirus exposed to a precise window of UV fluence reproducibly generates unique “non-replicating rhabdovirus-derived particles” (NRRPs) that maintain cell-entry and cytopathic properties. A gamut of leukemia cells, including multidrug-resistant blasts, underwent rapid NRRPs-induced apoptosis. Normal cell lines and healthy bone marrow mononuclear cells were resistant, in part through interferon-mediated signaling responses. Administering NRRPs intravenously was curative in a murine acute leukemia model, versus uniform disease progression using maximal tolerated dose of replicating virus. Serum levels of an array of immunomodulatory cytokines were significantly elevated after injection of NRRPs. iLOV prepared with NRRPs protected recipients from otherwise lethal leukemia. Intracranial administration of NRRPs proved nonlethal as opposed to neurotoxic live vesiculovirus. Following treatment, neutralizing antibodies were diminished with NRRPs compared to replicating virus. Together, NRRPs exhibit enhanced therapeutic index over replication-competent vesiculovirus. Leukemocidal activity of NRRPs is exerted through a plurality of immune-related and direct cytotoxic effects. This novel approach now extends vesiculovirus-based therapeutics into upfront treatment for acute leukemia.

Human acute nonlymphoblastic or myelogenous leukemia (ANLL or AML) is a malignant disease of the bone marrow involving hemopoietic (blood-forming) cells of the myeloid lineage. ANLL is a complex neoplastic disease, whose fundamental nature is only partially understood despite intensive research. The disease is complicated by its apparent heterogeneity in terms of the degree of differentiation of hemopoietic stem cell involvement in different patients and the cellular expression of immunologically defined surface markers. The presence of a common antigen in myelogenous leukemia (CAMAL) has been previously identified. This thesis examines the expression of the CAMAL marker in or on bone marrow (BM) and peripheral blood (PB) cells using a monoclonal antibody-based indirect immunoperoxidase slide test. Increased numbers of CAMAL-positive cells were found in or on BM and PB of myeloid leukemia patients (with acute or chronic forms of the disease) compared with those found in normals or most lymphoid malignancies. Results presented herein have demonstrated that fluctuations in CAMAL BM values (% positive cells) correlated with survival time prior to relapse. In a blind study, ANLL patients Whose CAMAL BM values decreased post-chemotherapy had significantly (p < 0.025) longer first remission times (x = 19.2 months) than patients with increasing or static CAMAL BM values (x = 6.8 months). CAMAL BM values were often observed to increase during remission, prior to relapse, suggesting the presence of residual subclinical disease. Addition of excess purified leukemia-derived CAMAL to an in vitro myeloid progenitor cell assay caused profound inhibition of normal CFU-c growth but had no inhibitory effect on CFU-c growth from myeloid leukemia patients in active disease states. Depletion of CAHAL from normal plasma and conditioned media (sources of numerous hemopoietic growth regulatory factors) caused significant inhibition of normal, but not myeloid leukemic, CFU-c growth. These results indicated that myeloid leukemic cells possessed apparent differences in responsiveness to CAMAL-mediated hemopoietic regulation compared to normal cells. Lack of responsiveness to inhibition by leukemia-derived CAMAL may facilitate dominance of the malignant clone over normal cells. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate

Acute Myeloid Leukemia (AML) is a type of cancer that affects the process of hematopoiesis. In individuals affected with AML, normal blood cells do not develop into red blood cells, white blood cells, and platelets, leading to symptoms such as anemia, neutropenia, and thrombocytopenia. The prognosis of AML is affected by multiple factors including: the genetic make-up of the leukemic cells, age of the affected individual, and underlying blood disorders such as myelodysplastic syndrome (MDS). MDS affects the development of stem cells into red blood cells, white blood cells, and platelets. Due to their clinical heterogeneity, AML and MDS continue to be a challenge that should be investigated in the community hospital setting. Remission rates between patients diagnosed with de-novo AML and patients diagnosed with AML with MDS were compared in a community hospital setting following induction therapy using a retrospective study design. Length of stay between patients diagnosed with de-novo AML and patients diagnosed with AML with MDS was compared during induction therapy. The association of age at diagnosis and number of chromosomal abnormalities to remission status was evaluated in each disease group. The association of blood transfusion requirements and neutropenic fever to length of stay was evaluated in each disease group. There were no statistically significant differences found between disease groups with respect to remission rates and length of stay. There were no statistically significant associations found between blood transfusion requirements and neutropenic fever in each disease group. There was an association found between age at diagnosis and remission status in patients diagnosed with AML with MDS. This indicates that older patients with AML with MDS are less likely to benefit from therapy and achieve complete remission. It is important to consider the small sample size, rare nature of the disease, and other variables that could have contributed to trends seen in the study population. The impact of predictors such as growth factor use and incidence of fungal infections should be investigated in future studies with AML patients. Considering these factors will allow for the development of targeted therapies and mechanisms against drug resistance for affected individuals.

La leucémie lymphoïde chronique (LLC) est décrite comme la plus fréquente forme de leucémie de l’adulte dans les pays occidentaux. Elle est caractérisée par l’envahissement de la moelle osseuse, du sang et parfois des organes lymphoïdes secondaires par des lymphocytes B matures à l’immunophénotype spécifique. Son évolution clinique est très variable selon les patients. Des progrès thérapeutiques majeurs permettent des réponses profondes et durables sans pour autant être à l’origine d’authentiques guérisons. Nous proposons dans ce travail d’explorer différents mécanismes de chimiorésistance. L’altération de TP53 est le facteur le plus reconnu de résistance mais les mécanismes qui lui sont possiblement associés sont peu documentés. Une première partie portant sur 115 patients a décrit une perte d’intégrité des télomères significativement plus importante en cas d’altération de TP53 (raccourcissement télomérique, ré-expression d’hTERT, répression des gènes codant le complexe shelterin). Ces 2 perturbations pouvant coopérer en aggravant l’instabilité génomique comme en témoignent des données cytogénétiques. Une autre analyse par séquençage ciblé nouvelle génération de 114 patients présentant une LLC en rechute a révélé la présence et le pronostic défavorable des combinaisons récurrentes de mutations géniques impliquant TP53, SF3B1 et ATM, gènes disposant d’un rôle crucial dans la réponse aux dommages à l’ADN. Une seconde partie s’est intéressée au rôle de TCL1A, une protéine co-activatrice d’AKT et de la voie NF-Kappa B. Alors que son hyperexpression est à l’origine du principal modèle murin de LLC, son rôle chez les patients restait peu connu. Nous avons montré qu’un niveau élevé d’expression de TCL1A était associé à des caractéristiques clinico-biologiques de maladie agressive et à un pronostic plus sombre. Le transcriptome dépendant de TCL1A généré chez des patients et dans une lignée lymphoïde B par une stratégie de shRNA est en faveur d’une signature MYC. Les inhibiteurs des protéines à BET bromodomaine ont précédemment été identifiés comme une stratégie efficace pour cibler cette voie. Cette approche thérapeutique (molécule JQ1) s’est avérée très efficace in vitro sur des cellules primaires de LLC.Ces travaux ont ainsi identifié des coopérations génomiques pouvant être à l’origine d’une résistance à l’immunochimiothérapie et ont aussi montré qu’une hyperexpression de TCL1A pouvait expliquer certains cas de résistance pour lesquels une stratégie de traitement épigénétique doit être évaluée dans des essais cliniques. / Résumé indisponible.

Les leucémies aigues myéloïdes (LAM) sont caractérisées par une expansion clonale de cellule(s) souche(s) leucémique(s) bloquée à un stade précoce de maturation. Malgré les avancées thérapeutiques, leur pronostic reste sombre et des progrès thérapeutiques doivent encore être réalisés. Dans les LAM, les espèces réactives de l’oxygène (ROS) sont considérées comme, d’une part, participant à la leucémogenèse et, d’autre part, comme hautement impliquées dans la sensibilité aux chimiothérapies conventionnelles. Par ailleurs, l’équilibre redox qui participe aux dérégulations métaboliques associées au processus leucémique, dépend de nombreux régulateurs, dont la protéine S100A8, protéine connue pour son action stimulatrice sur la NADPH oxydase et sa valeur pronostique dans les LAM.Ce travail s’est donc intéressé à la caractérisation des désordres oxydatifs dans les LAM afin d’évaluer leur impact clinico-biologique, et d’autre part au rôle de la sécrétion de la S100A8 dans le microenvironnement médullaire. De plus, à partir de lignées leucémiques, nous avons étudié l’impact de la S100A8 exogène sur la production de ROS, la respiration mitochondriale et le métabolome des cellules blastiques.A partir d’une cohorte de 84 patients atteints de LAM de novo au diagnostic, nous avons mis en évidence des désordres de l’équilibre redox à la fois dans les cellules leucémiques, dans les cellules normales de l’environnement médullaire ainsi que sur les systèmes régulateurs antioxydants (SOD, GPX, glutathion…). De plus, nous avons montré que la production des ROS observée en réponse à des modulateurs de la mitochondrie, qui reflète indirectement la fonctionnalité mitochondriale, joue un rôle pronostique indépendant des facteurs pronostiques habituels. L’analyse de la S100A8 dans les plasmas médullaires montre une expression augmentée dans les LAM, d’origine monocytaire majoritairement et est associée à des anomalies moléculaires de bon pronostic (inv(16), NPM1) ou un sous-groupe de patients FLT3-ITD mutés présentant une meilleure survie. Enfin, l’étude de la S100A8 sur les lignées leucémiques a permis de mettre en évidence la diversité de ses effets sur la croissance cellulaire, l’apoptose, la production de ROS ainsi qu’une variation métabolique de la phosphocholine dont les mécanismes restent à explorer.En conclusion, mon travail apporte des éléments originaux sur les particularités de l’équilibre bio-énergétique dans les LAM. Il souligne, que l’impact de ses dérégulations sur le pronostic des patients résulte de la combinaison d’un ensemble de facteurs métaboliques, qui doivent être appréhender dans leur globalité pour une meilleure efficacité thérapeutique. / Acute myeloid leukemia (AML) is characterized by clonal expansion of leukemic(s) cell(s) blocked at an early stage of maturation. Despite therapeutic advances, their prognosis remains poor and therapeutic improvements are needed. In AML, reactive oxygen species (ROS) are considered to contribute to leukemogenesis and, on the opposite, standard chemotherapies exert cytotoxicity via ROS. In addition, the redox balance acts on metabolic dysregulation in AML and depends on many regulators, such as S100A8 protein, associated with worst prognostic in AML and known to stimulate NADPH oxidase.In this context, this work focuses on oxidative disorders, and S100A8 expression in bone marrow microenvironment according to clinical-biological characteristics and evaluate their prognostic impact in AML. In addition, we investigated the impact of exogenous S100A8 on ROS production, mitochondrial respiration, and metabolism in leukemia cell lines.In a cohort of 84 de novo AML at diagnosis, we demonstrate the existence of redox balance disorders on leukemic cells, on normal cells from bone marrow microenvironment, and on antioxidant systems (SOD, GPX, glutathione ...). In addition, ROS production observed in response to mitochondrial modulators indirectly reflects mitochondrial functionality plays a prognostic role independent of the current prognostic factors. The analysis of S100A8 in bone marrow plasmas shows a higher expression in AML than in healthy controls or other hematological neoplasms. This hyperexpression is predominantly of monocytic origin and is associated with molecular abnormalities of good prognosis such as (inv (16), NPM1) or with a subgroup of mutated FLT3-ITD patients with better survival. Finally, the study of S100A8 on leukemia cell lines highlights its heterogeneous effect on cell growth, apoptosis, ROS production and on NOX regulation. Furthermore, we observe a S100A8-phosphocholine change which remains to be explored.In conclusion, this work provides original information on bio-energetic balance in AML and their prognostic impacts, emphasizing that these metabolic alterations impact AML prognosis through complex interactions.