Дисертації з теми "B-cell malignancie"
Оформте джерело за APA, MLA, Chicago, Harvard та іншими стилями
Ознайомтеся з топ-50 дисертацій для дослідження на тему "B-cell malignancie".
Біля кожної праці в переліку літератури доступна кнопка «Додати до бібліографії». Скористайтеся нею – і ми автоматично оформимо бібліографічне посилання на обрану працю в потрібному вам стилі цитування: APA, MLA, «Гарвард», «Чикаго», «Ванкувер» тощо.
Також ви можете завантажити повний текст наукової публікації у форматі «.pdf» та прочитати онлайн анотацію до роботи, якщо відповідні параметри наявні в метаданих.
Переглядайте дисертації для різних дисциплін та оформлюйте правильно вашу бібліографію.
Runarsson, Gudmundur. "Biosynthesis of leukotriene B₄ in hematological malignancies /." Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-386-8/.
Повний текст джерелаGreen, Michael R. "Molecular Profiling of B-Cell Malignancies." Thesis, Griffith University, 2009. http://hdl.handle.net/10072/366546.
Повний текст джерелаThesis (PhD Doctorate)
Doctor of Philosophy (PhD)
School of Medical Science
Griffith Health
Full Text
Mosti, Laura [Verfasser], and Anton [Akademischer Betreuer] Cathomen. "Generation of safe CAR T cells to target B cell malignancies." Freiburg : Universität, 2021. http://d-nb.info/1232174378/34.
Повний текст джерелаKokhaei, Parviz. "Preclinical therapeutic vaccination strategies in malignancies with focus on B-cell chronic lymphocytic leukemia /." Stockholm, 2006. http://diss.kib.ki.se/2006/91-7140-595-X/.
Повний текст джерелаMartínez-Martín, Sandra. "Targeting MYC in B-cell haematologic malignancies." Doctoral thesis, Universitat Autònoma de Barcelona, 2020. http://hdl.handle.net/10803/670653.
Повний текст джерелаLa importancia de la función de MYC en el cáncer (y el origen del nombre de la oncoproteína) se descubrió a finales de los años 70, con la identificación de la secuencia del retrovirus aviar causante de la leucemia mielocítica. Durante más de 40 años de investigación, se ha subrayado la relevancia de esta proteína en la división celular normal y su implicación en la transformación tumoral. De hecho, una de las primeras conexiones entre la sobreexpresión de proto-oncogenes (como MYC), reordenamientos génicos y el cáncer se hizo en el linfoma de Burkitt, la leucemia mieloide crónica y los plasmacitomas en ratón. Teniendo en cuenta el papel que desempeña MYC en los cánceres, parece obvia la necesidad de desarrollar estrategias terapéuticas contra esta proteína. Sin embargo, dirigir terapias contra MYC era y sigue siendo, un reto, dadas las propiedades únicas que lo caracterizan: su carencia de estructura tridimensional, localización nuclear y ausencia de un "bolsillo" enzimático. A pesar de estas particularidades, muchos estudios han demostrado el impacto terapéutico potencial que tendría la inhibición de MYC, ya sea directa o indirecta. En esta tesis, describimos el potencial de la inhibición directa de MYC en el linfoma de Burkitt (BL) y el mieloma múltiple (MM), usando dos estrategias distintas: pequeñas moléculas peptidomiméticas y disruptores del complejo MYC/MAX/ADN (la mini-proteína Omomyc y una variante derivada, llamada variante 26 o V26): - La validación del potencial terapéutico de los peptidomiméticos se hizo en colaboración con una "startup" biotecnológica. A pesar de la prometedora eficacia demostrada in vitro, los compuestos provocaron toxicidad severa local in vivo, además de cambios en el comportamiento de los animales, que nos llevaron a discontinuar la investigación. - Respecto a la validación de la mini-proteína Omomyc como estrategia farmacológica para tratar el BL y el MM, aquí mostramos su eficacia in vitro y resultados preliminares en un modelo de pez cebra, donde el pretratamiento con Omomyc previene la colonización de la médula ósea. De esta forma, se evidencia por primera vez el uso potencial de este "candidato a fármaco" para el tratamiento de tumores líquidos. Nuestros resultados in vivo en ratones muestran que, aunque la administración de Omomyc como monoterapia tiene una eficacia limitada (probablemente debido a la insuficiente llegada de péptido a las células de BL y MM), su combinación con un inhibidor del proteasoma (terapia estándar del mieloma) resulta en efectos sinérgicos tanto in vitro como in vivo en modelos de mieloma. Además, hemos mostrado que la administración intravenosa de Omomyc encapsulada en liposomas es segura y que dicha formulación liposomal prolonga la vida media de Omomyc en el suero, aunque no conseguimos aumentar la entrada en las células diana. - Por último, hemos caracterizado la V26, un derivado de la mini-proteína Omomyc, diseñada con el objetivo de mejorar la localización nuclear y el escape de los endosomas. Aquí mostramos que, como Omomyc, la V26 puede homodimerizar y heterodimerizar con MAX, además de unirse al ADN en cualquiera de las dos formas diméricas. Como esperábamos, la V26 se mostró más nuclear e indujo muerte celular in vitro. Sin embargo, también resultó ser menos soluble que Omomyc, de forma que sería necesario hacer mejoras en su formulación para poder usarla in vivo. En conjunto, nuestros resultados sugieren que la mini-proteína Omomyc, u otros derivados, como la V26, pueden servir como base para el diseño de nuevos fármacos anti-MYC para el tratamiento del BL y MM. Además, la combinación de éstos con las terapias estándar podrían constituir una estrategia prometedora, mientras que la encapsulación en liposomas podría ayudar a resolver aquellos posibles problemas de biodisponibilidad derivados de su uso in vivo.
The importance of MYC function in cancer (and the origin of the oncoprotein's name) was discovered in the late '70s when the sequence of the avian retrovirus that causes myelocytic leukaemia was identified. Since then, over 40 years of unceasing research have highlighted the significance of this protein in regular cell division, and importantly, its involvement in malignant transformation. Indeed, some of the earliest connections between the higher expression of proto-oncogenes (such as MYC), genetic rearrangements and their relation to cancer development were made in Burkitt lymphoma, chronic myeloid leukaemia and mouse plasmacytomas. Given the role of MYC in cancer, the need for the design of therapeutic strategies against it seems obvious. However, targeting MYC was - and somehow, still is - challenging due to its unique properties: lack of defined three-dimensional, structure nuclear localisation and absence of enzymatic pocket. Despite these difficulties, many studies have shown the potential therapeutic impact of direct or indirect MYC inhibition. In this thesis, we outline the potential of direct MYC inhibition in Burkitt lymphoma (BL) and multiple myeloma (MM) making use of 3 different strategies: small molecule peptidomimetics and disruptors of the MYC/MAX/DNA complex (Omomyc mini-protein and the derivative variant 26 or V26): - In the first case, the validation of the peptidomimetics therapeutic potential was done in collaboration with a start-up biotech company. Despite evidencing some promising efficacy in vitro, the compounds displayed severe local toxicity in vivo, accompanied by changes in animal behaviour that prompted us to discontinue the investigation. - Regarding the validation of Omomyc mini-protein as a pharmacological approach in the treatment of BL and MM, we demonstrated in vitro efficacy and showed preliminary results in the prevention of bone marrow homing upon Omomyc pre-treatment in a zebrafish model, indicating, for the first time, the potential use of this drug candidate to treat liquid tumours. Our in vivo data in mice show that, even if the administration of Omomyc as monotherapy has limited efficacy (probably due to insufficient delivery of peptide to BL and MM target cells), the combination with a proteasome inhibitor (the standard of care for myeloma) both in vitro and in vivo displays synergic effects in myeloma models. In addition, we were able to show that intravenous administration of the Omomyc mini-protein encapsulated in liposomes was safe and the liposomal formulation prolonged the serum half-life of the mini-protein, although it did not promote increased penetrance in MM target cells. - Lastly, we characterised V26, a rationally designed derivative of the Omomyc mini-protein, meant to display improved nuclear localisation and endosomal escape. Here we evidenced that, like Omomyc, V26 can homodimerise and heterodimerise with MAX, as well as bind DNA in both dimeric forms. As expected, V26 displayed better nuclear localisation and induced cell death in vitro. However, it also turned out to be less soluble than Omomyc, indicating that it would require further formulation efforts to be used in vivo. Altogether, our results suggest that Omomyc mini-protein itself or other Omomyc-derivatives, like V26, can serve as the backbone for the design of new anti-MYC agents to treat BL and MM. In this context, combination therapy with the standard of care seems to be a promising strategy, while encapsulation in liposomes might help to address potential bioavailability issues that might arise from its use in vivo.
McCann, Katy. "Immunogenetic analysis of aggressive B-cell malignancies." Thesis, University of Southampton, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.494386.
Повний текст джерелаGupta, Sneha Veeraraghavan. "Targeting Protein Metabolism in B-cell Malignancies." The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1343169973.
Повний текст джерелаJiménez, Bernal Isabel. "Tumor immune microenvironment in B-cell lymphoid malignancies." Doctoral thesis, Universitat Autònoma de Barcelona, 2020. http://hdl.handle.net/10803/671173.
Повний текст джерелаEl microambiente inmune tumoral juega un papel fundamental en las etapas tempranas de la formación de los tumores y en la progresión de éstos. Terapias dirigidas a este microambiente ofrecen nuevas opciones terapéuticas y también sirven para mejorar las terapias actuales frente a muchos cánceres, incluyendo los que afectan a las células B. Sin embargo, son necesarias más investigaciones para entender en mayor profundidad los mecanismos de evasión del sistema inmune que favorecen la progresión de los tumores y diseñar inmunoterapias más precisas. Nuestros principales objetivos son aportar nuevas evidencias sobre mecanismos inmunes asociados a la progresión tumoral y las bases pre-clínicas para el desarrollo de nuevas estrategias terapéuticas con potencial inmuno-modulador. Para ello, nos centramos en la leucemia linfática crónica (LLC) y en el linfoma cerebral primario (LCP). Los mecanismos de progresión en LLC desde estadios tempranos no son conocidos en su totalidad. Aunque la adquisición de alteraciones moleculares es escasa sugiriendo que la LLC no progresa exclusivamente por mecanismos de evolución clonal, todavía no se ha llevado a cabo un análisis exhaustivo del microambiente inmune que demuestre que la progresión sí pueda deberse a cambios inmunes. Por ello, hemos realizado un estudio longitudinal abarcando tanto los escenarios genéticos como inmunológicos en pacientes de LLC sin tratar que han progresado clínicamente y en pacientes asintomáticos durante un largo periodo de tiempo. Nuestros resultados muestran que los pacientes que progresan experimentan un incremento de células T CD8+ efectoras de memoria y terminalmente exhaustas T-betmid/-EomeshiPDhi a la progresión. Este incremento no se observa en los pacientes de LLC que no han progresado. Además, las células T a la progresión adquieren un perfil transcripcional diferente. Esto va acompañado de un aumento en las propiedades inmunosupresoras de las células leucémicas a la progresión. Demostramos que las células de LLC en el momento de la progresión tienen mayor capacidad de inducir exhaustión tanto en células T CD8+ de LLC como aquellas procedentes de individuos sanos, y que lo hacen mediante un mecanismo dependiente de factores solubles que incluye IL-10. Los escasos cambios genéticos que encontramos tras secuenciar el exoma de nuestros pacientes nos permiten concluir que las variaciones inmunes que hemos identificado son fundamentales para la progresión de la LLC. El desenlace de los pacientes diagnosticados con LCP es normalmente desfavorable debido a la escasez de opciones terapéuticas efectivas. Las células malignas de LCP presentan con frecuencia una desregulación de la vía del receptor de la célula B (del inglés, BCR), pero su inhibición mediante ibrutinib muestra respuestas muy breves en pacientes. Sin embargo, la vía del BCR también puede bloquearse mediante la inhibición de la exportina nuclear XPO1 con selinexor. Selinexor atraviesa la barrera hemato-encefálica y ha mostrado actividad en un paciente diagnosticado con linfoma difuso de células grandes B con recaída en el sistema nervioso central. Por consiguiente, decidimos evaluar los efectos de selinexor en monoterapia y combinado con ibrutinib en modelos pre-clínicos murinos de LCP. Nuestro análisis muestra que selinexor bloquea el crecimiento tumoral y prolonga la supervivencia en un modelo de ratón bioluminiscente y la combinación con ibrutinib prolonga aún más la supervivencia. Demostramos que los linfomas cerebrales en ratón están infiltrados con macrófagos pro-tumorales M2 que expresan PD-1 y SIRPα. Además, el tratamiento con selinexor e ibrutinib favorece la respuesta inmune anti-tumoral induciendo un cambio en la polarización de los macrófagos hacia un perfil pro-inflamatorio y reduciendo la expresión de PD-1 y SIRPα en los macrófagos M2 asociados al tumor.
The tumor immune microenvironment (TIME) plays a critical role in the early formation of tumors and their progression. Targeting the TIME has offered new therapeutic approaches and improved current ones in several cancers, including B-cell malignancies. Nonetheless, further investigation is needed in order to more deeply understand immune evasion mechanisms that lead to tumor progression and to design therapies that modulate the immune system more precisely. Here, our main objectives are to provide new insights into immune mechanisms that favor tumor progression and a pre-clinical rationale for the design of new therapeutic strategies with immunomodulatory potential. To accomplish these goals our study will focus on chronic lymphocytic leukemia (CLL) and primary central nervous system lymphoma (PCNSL). Mechanisms driving the progression of CLL from its early stages are not fully understood. This hampers detecting progression in advance and developing therapies that could intervene in the early stages. Although the limited acquisition of molecular changes suggests that CLL progression is not mainly driven by clonal evolution, a deeper analysis of the immune microenvironment that demonstrates immune variations over time that contribute to progression has not been performed. Hence, we longitudinally studied the immune and genetic landscapes of untreated progressing and non-progressing patients. Our results show that progressed CLL patients experience an increase in effector memory and terminally exhausted T-betmid/-EomeshiPDhi CD8+ T cells over time, not observed in non-progressing patients. In addition, T cells at progression acquire a distinct transcriptional profile. This is accompanied by enhanced immunosuppressive properties in leukemic cells at progression. We prove that progressed CLL cells are intrinsically more capable of inducing CD8+ T-cell exhaustion in T cells affected by CLL and healthy T cells by a mechanism dependent on soluble factors including IL-10. In addition, the reduced genetic changes we found by whole-exome sequencing in our cohort indicate these immune variations are fundamental for progression in CLL. Patients diagnosed with PCNSL often face dismal outcomes due to the limited availability of therapeutic options. PCNSL cells frequently have deregulated B-cell receptor (BCR) signaling, but its inhibition using ibrutinib only offers a brief effective response in PCNSL patients. Nonetheless, the BCR pathway can also be blocked by inhibiting the nuclear exportin XPO1 using selinexor. Selinexor is able to cross the blood–brain barrier and has shown positive clinical activity in a patient with refractory diffuse large B-cell lymphoma in the CNS. Accordingly, we evaluated the effects of selinexor alone and also combined it with ibrutinib in pre-clinical mouse models of PCNSL. Our analysis shows that selinexor blocks tumor growth and prolongs survival in a bioluminescent mouse model and its combination with ibrutinib further increases survival. We demonstrate that CNS lymphomas in mice are infiltrated by tumor-promoting M2-like macrophages expressing PD-1 and SIRPα. Moreover, the treatment with selinexor and ibrutinib favors an anti-tumoral immune response by shifting macrophage polarization toward an inflammatory phenotype and diminishing the expression of PD-1 and SIRPα in M2 tumor-associated macrophages.
Caeser, Rebecca. "Elucidating oncogenic mechanisms in human B cell malignancies." Thesis, University of Cambridge, 2018. https://www.repository.cam.ac.uk/handle/1810/285011.
Повний текст джерелаForster, Jade. "Evaluating the genomic landscape of B cell malignancis." Thesis, University of Southampton, 2016. https://eprints.soton.ac.uk/408724/.
Повний текст джерелаPocock, Christopher Francis Elliot. "Modelling and molecular manipulation of human B cell malignancies." Thesis, Queen Mary, University of London, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.261876.
Повний текст джерелаAlcaraz, Amor. "ZFP36 proteins and mRNA targets in B cell malignancies." Thesis, University of Westminster, 2015. https://westminsterresearch.westminster.ac.uk/item/q254z/zfp36-proteins-and-mrna-targets-in-b-cell-malignancies.
Повний текст джерелаLim, Sean H. "Investigation of CD20-directed immunotherapy in B-cell malignancies." Thesis, University of Southampton, 2011. https://eprints.soton.ac.uk/376893/.
Повний текст джерелаRafiq, Sarwish. "Evaluation of Antibody-based Therapeutics in B cell Malignancies." The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1338321515.
Повний текст джерелаKarlsson, Hannah. "CD19-targeting CAR T Cells for Treatment of B Cell Malignancies : From Bench to Bedside." Doctoral thesis, Uppsala universitet, Klinisk immunologi, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-232638.
Повний текст джерелаDubosq, Ming-Celine. "The non-viral production of Chimeric Antigen Receptor T-cells for B-cell haematological malignancies." Thesis, University of Sydney, 2020. https://hdl.handle.net/2123/23170.
Повний текст джерелаMouzakiti, Amalia. "Regulation of death receptor-mediated apoptosis in B cell malignancies." Thesis, University of Southampton, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.271649.
Повний текст джерелаWalter, Harriet Sarah. "Studies of Bruton's tyrosine kinase inhibitors in B-cell malignancies." Thesis, University of Leicester, 2018. http://hdl.handle.net/2381/42887.
Повний текст джерелаSIMONETTI, GIORGIA. "B lymphoid malignancies: insights from mouse models." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2012. http://hdl.handle.net/10281/30033.
Повний текст джерелаBrown, Alison G. "Molecular analysis of 13ql4 abnormalities in B-cell malignancy." Thesis, University of Edinburgh, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.306244.
Повний текст джерелаFlordal, Thelander Emma. "Genetic characterization of hematological malignancies with focul on mantle cell lymphoma /." Stockholm, 2007. http://diss.kib.ki.se/2007/978-91-7357-161-6/.
Повний текст джерелаSamuel, Jesvin John. "Translational studies in B-cell malignancies : studies on TP53 and BRAF." Thesis, University of Leicester, 2015. http://hdl.handle.net/2381/36233.
Повний текст джерелаIsraelsen, Nathan. "Surface-Enhanced Raman Spectroscopy-Based Biomarker Detection for B-Cell Malignancies." DigitalCommons@USU, 2015. https://digitalcommons.usu.edu/etd/4605.
Повний текст джерелаMcWilliams, Emily Mary. "Restoring Innate NK-cell Immunity with Antibody Therapeutics in CLL B-Cell Malignancy." The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1479863842166353.
Повний текст джерелаBishop, David C. "Clinical Translation of CD19-Specific Chimeric Antigen Receptor T cells Generated with the piggyBac Transposon System for the Treatment of B Cell Malignancies." Thesis, The University of Sydney, 2019. https://hdl.handle.net/2123/22305.
Повний текст джерелаMcCarthy, Helen. "Immunogenetic analysis of B-cell malignancies and the clinical application of DNA idiotypic vaccines." Thesis, University of Southampton, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.418059.
Повний текст джерелаPark, Eugene. "Chemosensitisation of B-cell malignancies through inhibition of microenvironmental Protein Kinase C-beta (PKCβ)". Thesis, University of Cambridge, 2019. https://www.repository.cam.ac.uk/handle/1810/288230.
Повний текст джерелаCapolla, Sara. "Use of immune-nanoparticles containig chemiotherapeutic agents for the treatment of B-cell malignancies." Doctoral thesis, Università degli studi di Trieste, 2015. http://hdl.handle.net/10077/10980.
Повний текст джерелаB-cell malignancies are a heterogeneous group of clinical conditions including indolent diseases such as Chronic Lymphocytic Leukemia (CLL) and highly aggressive lymphoproliferative disorders such as Burkitt’s lymphoma. B-cell malignancies treatments take advantage of dose-intensive chemotherapeutic regimens and immunotherapy via monoclonal antibodies. Unfortunately, they may lead to insufficient tumor distribution of therapeutic agents and cause several adverse effects. Thus, we propose a novel therapeutic approach for the treatment of CLL and Burkitt’s lymphoma in which high-doses of the association of hydroxychloroquine and chlorambucil (HCQ/CLB) or fludarabine were loaded inside biodegradable nanoparticles (BNPs) coated with an anti-CD20 antibody. First of all, a Burkitt’s lymphoma cell line (BJAB), two CLL cell lines (MEC1 and EHEB) and cells purified from patients’ blood samples were used to confirm CD20 expression and to assess BNPs binding and internalization. These studies demonstrated BNPs ability to bind malignant B cells and to enter inside cells in a process different from endocytosis. Then, BNPs therapeutic effect was evaluated by MTT test, AnnV/PI assay and western blot to put in evidence apoptosis induction and autophagy inhibition. These experiments demonstrated drugs-loaded BNPs ability to kill malignant B cells with comparable effects than those obtained with free drugs whereas empty BNPs were practically ineffective. In vivo BNPs characterization included the evaluation of their toxicity, biodistribution and therapeutic effect. C57/BL mice were used to evaluate BNPs toxicity which was studied considering survival, loss of body weight and several tissue markers in the blood. Mice receiving 8 injections of free HCQ+CLB died in this experiment whereas animals challenged with the same amount of drugs encapsulated inside BNPs did not show toxic effects suggesting BNPs safety. The importance of antiCD20 antibody in the homing of BNPs was confirmed by in vivo Time-Domain Optical Imaging performed in localized B-cell malignancy-bearing mice. This analysis suggested the ability of antiCD20-conjugated BNPs to specifically target tumor B-cells, with a pick after 24-48 hours. On the contrary, untargeted BNPs localization inside tumor was significantly decreased. In this analysis it was also evident that the liver is the main site of BNPs’ elimination while in the other organs the presence of fluorescent BNPs was very low. Finally, BNPs ability to treat a new xenograft human/SCID leukemia and Burkitt’s lymphoma mouse model was studied. Drugs-loaded BNPs were able to improve HCQ/CLB efficacy in vivo allowing the cure of treated all Burkitt’s lymphoma-bearing mice and 3 out of 7 leukemia-bearing animals. All these data together put the basis for the potential use of BNPs in the treatment of B-cell malignancies.
I tumori a cellule B sono un gruppo eterogeneo di patologie che comprendono sia malattie indolenti, come la leucemia linfatica cronica (LLC), sia aggressive, come il linfoma di Burkitt. Il trattamento delle patologie a cellule B prevede sia l’utilizzo di chemioterapici (agenti alchilanti e analoghi delle purine) che di anticorpi monoclonali. Nonostante la varietà di terapie esistenti, l’efficacia di questi farmaci è limitata dalla mancata specificità per le cellule tumorali e dall’induzione di gravi effetti collaterali. Per ovviare ai limiti delle terapie attuali, è stato quindi proposto l’utilizzo di nanoparticelle coniugate con un anticorpo antiCD20, specifico per le cellule B, e contenenti alte concentrazioni di chemioterapici (idrossiclorochina e clorambucile o fludarabina). Le nanoparticelle sono state caratterizzate in vitro e in vivo durante questo progetto di dottorato. Inizialmente sono stati effettuati studi in vitro al fine di valutare l’espressione del CD20 sulla superficie di una linea cellulare di linforma di Burkitt (BJAB), due linee di LLC (MEC1 e EHEB) e cellule purificate da campioni di sangue di pazienti affetti da LLC. In seguito, il legame e l’internalizzazione delle nanoparticelle a queste cellule sono stati dimostrati suggerendo anche come le nanoparticelle vengano internalizzate attraverso un meccanismo diverso dall’endocitosi. L’effetto terapeutico in vitro delle nanoparticelle è stato valutato con test MTT, AnnessinaV/PI e tramite western blot al fine di evidenziare l’induzione di apoptosi e l’inibizione dell’autofagia, meccanismi attraverso cui i farmaci utilizzati sono noti agire. Questi esperimenti hanno dimostrato che le nanoparticelle cariche di chemioterapici sono in grado di uccidere le cellule B tumorali con effetti paragonabili a quelli ottenuti da pari concentrazioni di farmaci liberi dimostrando come il processo di produzione delle nanoparticelle non influisca sull’efficacia dei chemioterapici. Al contrario, nanoparticelle vuote non sono in grado di uccidere le cellule dimostrando la mancata tossicità dei polimeri da cui sono costituite. Dopo aver confermato il legame e l’internalizzazione delle nanoparticelle che inducono la morte delle cellule B tumorali, sono stati effettuati esperimenti in vivo tra cui studi di tossicità al fine di valutare eventuali effetti collaterali indotti dal trattamento, studi di biodistribuzione e la valutazione degli effetti terapeutici. Gli studi di tossicità sono stati effettuati in topi sani valutando parametri quali la perdita di peso, la sopravvivenza e la tossicità sistemica. Nanoparticelle cariche di farmaci presentano un profilo tossicologico sicuro mentre pari dosi di farmaci liberi inducono la morte di tutti gli animali trattati. Questi esperimenti dimostrano quindi come l’inserimento di farmaci all’interno di nanoparticelle prevenga gli effetti collaterali normalmente indotti dai chemioterapici. Secondariamente, sono stati effettuati studi di biodistribuzione di nanoparticelle coniugate o meno con un anticorpo antiCD20. Questi studi effettuati tramite Optical Imaging dimostrano come nanoparticelle coniugate con l’anticorpo antiCD20 si localizzino preferenzialmente nella massa tumorale in 24-48 ore in quantità maggiore rispetto a nanoparticelle non coniugate con l’anticorpo. Inoltre, da queste analisi risulta evidente come il fegato sia il maggiore sito di eliminazione delle nanoparticelle mentre in altri organi la presenza di nanoparticelle è molto bassa. Infine, un modello disseminato di linfoma di Burkitt e un modello di LLC sono stati sviluppati in topi SCID (Severe Combined ImmunoDeficiency) iniettando rispettivamente cellule BJAB intraperitoneo e cellule MEC1 endovena. I modelli sono stati caratterizzati e utilizzati per valutare la potenziale applicazione delle nanoparticelle nel trattamento di queste patologie. Questi studi hanno dimostrato come le nanoparticelle siano in grado di aumentare l’efficacia dei chemioterapici e di curare tutti i topi affetti da linfoma di Burkitt e 3/7 topi affetti da leucemia. Concludendo, questi risultati suggeriscono la potenziale applicazione delle nanoparticelle cariche di chemioterapici nel trattamento di LLC e linfoma di Burkitt.
XXVII Ciclo
1986
Rozmus, Jacob. "Novel mechanisms involving B cell receptor (BCR) and B cell activating factor (BAFF) signaling pathways underlying human primary immunodeficiencies and malignancy." Thesis, University of British Columbia, 2016. http://hdl.handle.net/2429/58377.
Повний текст джерелаMedicine, Faculty of
Pathology and Laboratory Medicine, Department of
Graduate
Lin, Bin Liang Kevin. "The role of the Rap GTPases in B-cell morphology, function, and malignancy." Thesis, University of British Columbia, 2009. http://hdl.handle.net/2429/9900.
Повний текст джерелаLaffan, Michael. "Anomalous gene rearrangements in B-cell malignancies : implications for the mechanism of immunoglobulin class switch." Thesis, University of Oxford, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.336094.
Повний текст джерелаFarren, Timothy william. "The role of the NK cell receptor CD160 in the diagnosis, differentiation and function of chronic B-cell malignancies." Thesis, Queen Mary, University of London, 2013. http://qmro.qmul.ac.uk/xmlui/handle/123456789/9011.
Повний текст джерелаLiu, Hui. "The molecular mechanisms of cyclin D1 gene deregulation by immunoglobulin regulatory sequences in B cell malignancies." Diss., The University of Arizona, 2004. http://hdl.handle.net/10150/280522.
Повний текст джерелаHaga, Christopher L. "Analysis of the role of FCRL5 and FIGLERs in B cell development, signaling and malignancy." Thesis, Birmingham, Ala. : University of Alabama at Birmingham, 2008. https://www.mhsl.uab.edu/dt/2008d/haga.pdf.
Повний текст джерелаHammarström, Viera. "B-cell immunity in patients with hematological malignancies and after stem cell transplantation : studies with special reference to tetanus and pneumococcal immunity /." Stockholm, 1998. http://diss.kib.ki.se/search/diss.se.cfm?19980828hamm.
Повний текст джерелаMani, Rajeswaran. "Preclinical development of a non-immunosuppressive FTY720 derivative OSU-2S forchronic lymphocytic leukemia and other B-cell malignancies." The Ohio State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=osu1404067069.
Повний текст джерелаWillis, Anthony Graham. "Rapid molecular cloning of immunoglobulin translocations in B-cell malignancy by long-distance inverse polymerase chain reaction." Thesis, King's College London (University of London), 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.312267.
Повний текст джерелаPIEVANI, ALICE SILVIA. "Cytokine-induced killer (cik) cell cultures for the adoptive immunotherapy of hematological malignancies: characterization and new therapeutic strategies for clinical application." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2011. http://hdl.handle.net/10281/20178.
Повний текст джерелаDey, A. "Enhancing the neutrophil-mediated anti-cancer response after oncolytic measles virus therapy in B cell malignancy : dissecting out the mechanism." Thesis, University College London (University of London), 2016. http://discovery.ucl.ac.uk/1521057/.
Повний текст джерелаLo, Yee-nga, and 盧懿雅. "Effect of t(11;14)(p13;q32) translocation on the expression of PDHX, the telomeric gene on chromosome 11p13, in mature B-cell malignancies." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2011. http://hub.hku.hk/bib/B46632505.
Повний текст джерелаChoudhary, Gaurav Sudhakar. "Role of Myeloid Cell Leukemia 1 (MCL-1) in mediating chemoresistance towards BCL-2 homology 3 (BH3) mimetics in lymphoid malignancies." Case Western Reserve University School of Graduate Studies / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=case1448024862.
Повний текст джерелаOuedraogo, David Eric. "Exploration du réservoir EBV chez les patients infectés par le VIH : implications pathologiques." Thesis, Montpellier 1, 2013. http://www.theses.fr/2013MON1T001.
Повний текст джерелаIt is assumed that circulating memory B cells including those latently infected by EBV return periodically to lymphoid nodes where they are stimulates and undergo differentiation into immunoglobulin-producing cells allowing the virus to initiate viral replication. However, the monitoring and the management of EBV reactivation and it association with lymphoid malignancies in HIV-infected patients are still being controversies and need a better understanding of the probable mechanisms involved. In this study, we proposed novel biological tools for EBV DNA quantification allowing discriminating latent and lytic reservoir. We showed that the EBV reservoir levels are closely associated with the polyclonal B-cell activation. We also observed an association between immune activation and EBV reactivation markers with the occurrence of B-cell lymphoma. Moreover, we described a long term evolution of monoclonal gammapathies in HIV-infected subjects and the persistence of the immunoglobulis monoclonal pike was found to be associated with higher EBV reservoir levels. Therefore, the B-cell activation and subsequently EBV reactivation likely play the main roles on the occurrence of B lymphocytes malignancies during HIV infection
Zangrossi, Manuela. "Study of the extra-telomeric functions of telomerase in in vitro and in vivo models." Doctoral thesis, Università degli studi di Padova, 2018. http://hdl.handle.net/11577/3426233.
Повний текст джерелаIl mantenimento dei telomeri, necessario per la proliferazione illimitata delle cellule tumorali, è esercitato dalla telomerasi, un complesso ribonucleoproteico contenente una trascrittasi inversa specializzata, codificata dal gene TERT, che utilizza un templato ad RNA per sintetizzare nuove sequenze telomeriche, svolgendo quindi un ruolo critico nella formazione e nella progressione dei tumori. TERT viene infatti solitamente represso in normali cellule somatiche, mentre è rilevabile nella maggior parte dei tumori. Studi recenti hanno suggerito che TERT è coinvolto in altre funzioni cellulari e può contribuire alla carcinogenesi anche attraverso meccanismi indipendenti dal mantenimento dei telomeri, quindi la sua inibizione potrebbe rappresentare una strategia promettente per migliorare il trattamento antitumorale, al di là dell’effetto sui telomeri. I possibili effetti terapeutici di BIBR1532 (BIBR), un inibitore specifico del TERT, sono stati valutati in diversi contesti cellulari, ma non sono attualmente disponibili dati ottenuti su modelli di neoplasie delle cellule B sia associate al virus di Epstein-Barr (EBV) che virus-indipendenti. Lo scopo di questo studio era di caratterizzare gli effetti biologici dell'inibizione di TERT a breve termine da parte del BIBR su linee cellulari linfoblastoidi immortalizzate da EBV (LCL) e su modelli in vitro di linfoma di Burkitt (BL); inoltre, sono stati studiati gli effetti del trattamento con BIBR a breve termine in vivo negli embrioni di zebrafish. I risultati ottenuti hanno dimostrato che l'inibizione a breve termine di TERT da parte di BIBR, in modelli in vitro di tumori delle cellule B, ha portato a una diminuzione della proliferazione cellulare, all'accumulo di cellule nella fase S e infine all'aumento dell'apoptosi. L'arresto del ciclo cellulare e l'apoptosi, conseguenti all'inibizione di TERT a breve termine, erano associati e probabilmente dipendenti dall'attivazione della risposta al danno del DNA, come evidenziato dall’aumento dei livelli di γH2AX e dall'attivazione dei pathway di ATM e ATR. L’analisi della media e del range di lunghezza dei telomeri e dei foci di danno al DNA ha indicato che la risposta al danno attivata in seguito all’inibizione TERT a breve termine non era legata a disfunzioni telomeriche, suggerendo quindi che TERT, oltre a stabilizzare il telomero, può proteggere il DNA tramite meccanismi telomero-indipendenti. In particolare, LCL-TERT positive trattate con BIBR in combinazione con fludarabina o ciclofosfamide hanno mostrato un aumento significativo del numero di cellule apoptotiche rispetto a quelle trattate con agenti chemioterapici da soli. In accordo con i risultati in vitro, l'inibizione a breve termine di Tert da parte del BIBR in embrioni di zebrafish ha ridotto la proliferazione cellulare, indotto un accumulo di cellule nella fase S, aumentato il tasso di apoptosi e innescato l'attivazione della risposta al danno al DNA. Questi effetti non erano legati a disfunzioni telomeriche, poiché il range di lunghezza dei telomeri non era influenzato dal trattamento a breve termine con BIBR e i foci di danno al DNA erano distribuiti casualmente, piuttosto che localizzati in modo specifico sui telomeri. Tutti questi effetti erano specificamente associati all'inibizione di Tert poiché il trattamento con BIBR non mostrava alcun effetto negli embrioni di zebrafish Tert-negativi. Nel complesso questi dati dimostrano che l'inibizione del TERT compromette la proliferazione cellulare e induce effetti pro-apoptotici non associati a disfunzioni telomeriche, rafforzando il concetto che TERT esercita di per sé funzioni pro-tumorali indipendenti dalla lunghezza del telomero e quindi supportando l'introduzione di inibitori di TERT per integrare le attuali modalità di trattamento antitumorale.
Dandoit, Mylène. "Evaluation de l'impact de la prise en charge thérapeutique sur la survie et la qualité de vie des patients atteints d'un lymphome folliculaire ou d'un lymphome B diffus à grandes cellules." Thesis, Dijon, 2014. http://www.theses.fr/2014DIJOS038/document.
Повний текст джерелаIn France, hematologic malignancies, which are the sixthmost common cancers, are amajor public healthproblem. This work aimed to study the impact of the therapeutic management on survival and healt-relatedquality of life (HRQoL) in patients with these hematologic malignancies. The first objective of this work is topresent an overview of the epidemiology of lymphoid malignancies with a study of changes in the incidenceand net survival in the Côte d’Or department between 1980 and 2009. The incidence, which has increased since1980, seems to have stabilized since the 2000s for some entities, including follicular lymphoma (FL) and diffuselarge B-cell lymphoma (DLBCL). Overall, we observed an improvement in net survival, with, however, a lessfavorable prognosis in the short and long-term for some entities. FL and DLBCL were the first lymphomas tobenefit from the introduction of monoclonal antibodies in their therapeutic management. Our second studyaimed to assess the impact of rituximab on overall survival in patients with FL or DLBCL in the Côte d’Or departmentusing a methodology based on the propensity score. Our results confirmed the significant benefit ofrituximab on overall survival in an unselected population of patients. In view of these results, we studied theHRQoL of these patients during and after treatment. HRQoL evolved differently during follow-up dependingon the type of lymphoma
Koduri, Megha Pallavi. "The curative potential of chimeric antigen receptor T-cell therapy for B-cell malignancies." Thesis, 2017. https://hdl.handle.net/2144/23818.
Повний текст джерелаLiu, Hui. "Functional studies of BCL11A a transcriptional repressor implicated in chromosome 2p13-disrupted malignancy /." Thesis, 2002. http://wwwlib.umi.com/cr/utexas/fullcit?p3099485.
Повний текст джерелаMaswabi, Bokang Calvin. "Charakterizace hematopoetických buněk u pacientů s nádorem ze zralých B buněk." Doctoral thesis, 2017. http://www.nusl.cz/ntk/nusl-372368.
Повний текст джерелаBerry, Nadine Kaye. "Clinical use of SNP-microarrays for the detection of genome-wide changes in haematological malignancies with a focus on B-cell neoplasms." Thesis, 2020. http://hdl.handle.net/1959.13/1412536.
Повний текст джерелаHaematological malignancies are a heterogenous group of cancers that originate in the mature or immature cells of the haemopoietic system. Some originate in the bone marrow, where stem cells differentiate into many types of immature blood cells, while others may form in the peripheral blood once the blood cell has matured. Each type of haematological malignancy is usually characterised by a distinct set of genomic changes that, when identified, can be used to make a specific diagnosis, predict response to therapy, provide a risk for relapse or chance of survival, characterise the disease further or provide a therapeutic target. These genomic changes are often well defined and recurrent in each type of haematological malignancy. Recent advances in technologies have also enabled the rapid discovery of novel findings – some of which have not been detectable by current standard clinical investigations performed at diagnosis. There are several techniques used to investigate the genome of haematological malignancies including G-banded karyotype, fluorescent in situ hybridisation (FISH) and various sequencing technologies. Currently, the primary technique used for the clinical genomic investigation of haematological malignancies is the G-banded karyotype, whereby analysis of banding patterns on chromosomes, which are visible in the metaphase of a cell cycle, is performed using a light microscope. This method of analysis has been the cornerstone of haematological malignancy characterisation since the discovery of the Philadelphia (‘Ph’) chromosome in 1960. However, there are many known limitations of this technique with low resolution and the need for dividing malignant cells being the most obvious. New cytogenomic tools, such as comparative genomic hybridisation microarray (CGH-microarray) and single nucleotide polymorphism microarray (SNP-microarray), overcome both the limitation of low resolution and the need for dividing cells by enabling the investigation of the whole genome at high resolution using DNA. However, the uptake of these techniques into clinical practice has been slow for haematological malignancy investigations. In this research endeavour, I sought to evaluate the clinical use of microarray technology for the investigation of haematological malignancies by assessing the performance of both CGH and SNP microarrays in comparison to current techniques. I focused on the platform’s ability to detect genomic changes that are known to hold prognostic value, as well as their ability to improve the detection of novel changes and those which are not detectable by current methodologies. In the first part of this research project I undertook a detailed literature review. Subsequently, I investigated the clinical use of CGH-microarrays for the detection of copy number variants (CNV’s) in haematological malignancies. The findings were compared to those observed using the current standard clinical investigations followed by a discussion of its prognostic implications. In the next step I used SNP-microarrays to further characterise the CGH-microarray findings. I evaluated the SNP-microarray for the identification and classification of prognostically important complex genomic signatures, such as chromothripsis and chromoanasynthesis, which are not identifiable by current clinical investigative tools. Finally, the impact of the detection of new prognostically important CNV’s identified at diagnosis by SNP-microarray were explored in a specific disease cohort. The literature clearly supports the use of new genomic profiling techniques that are only now beginning to transition from the research setting into clinical practice. Experts in the field of haematological malignancies recognise the role of genomic profiling as important for the understanding of this disease subset. However, research is still required to determine how significant the findings are and how to personalise treatment for patients based on these findings. It is imperative that the clinical diagnostic and treatment regimens do not fall too far behind advances made in research, particularly when it can have an immediate and positive effect on the patient.
Bai, Li-Yuan, and 白禮源. "Treating B-cell lymphoid malignancies with non-conventional chemotherapeutic agents: study of compounds targeting p38 MAPK, Akt pathways or histone deacetylase." Thesis, 2011. http://ndltd.ncl.edu.tw/handle/00407222502301962749.
Повний текст джерела中國醫藥大學
臨床醫學研究所博士班
99
Hematological malignancies can be grouped as either myeloid or lymphoid disorders. The standard treatment strategy for lymphoid malignancies is chemotherapeutic agents, usually in combination with monoclonal antibodies which have introduced great impact on the clinical situation. Although certain subtypes of lymphoid malignancies, e.g. diffuse large B cell lymphoma, have good response to initial treatment, some of patients will succumb to the disease because of development of drug resistance. Still others are primarily refractory to initial therapy. The prognosis for these patients is dismal. This highlights the necessity of developing approaches to improve the therapeutic efficacy for lymphoid malignancies. One approach is to develop compounds that have anti-tumor effects with mechanism different from conventional chemotherapeutic agents. Another strategy to improve the therapeutic efficacy is to sensitize cancer cells to cytotoxic effect of other agents. We hypothesize that agents targeting alternative pathways are effective in treatment of lymphoid malignancies. These agents can exhibit anti-tumor efficacy either by themselves or by sensitizing cancer cells to other cytotoxic agents. For this sack, we validate our hypothesis through three approaches. The first part demonstrates that OSU-DY7 induces cytotoxicity in chronic lymphocytic leukemia and Burkitt lymphoma through activating p38 mitogen-activated protein kinase pathway. The second part illustrates the effectiveness of a histone deacetylase inhibitor (S)-HDAC42 in multiple myeloma cells U266. The last part shows that OSU-03012, an Akt pathway inhibitor, sensitizes myeloma cells to imatinib mesylate via AMP-activated protein kinase and STAT3 pathways. Our study successfully proves the hypothesis that agents targeting p38 MAPK, Akt pathways or histone deacetylase are effective in treatment of lymphoid malignancies. Further studies are needed to shed more light on the road.
André, Ana Filipa Santos. "Selection of single domain antibodies against the CD20 receptor: a new treatment with potencial anti-tumor properties for B-cell malignancies." Master's thesis, 2016. http://hdl.handle.net/10451/25211.
Повний текст джерелаLymphoma is the third most common neoplasia in the world. Within lymphomas, non-Hodgkin lymphoma (NHL) is the most common affecting mainly B cells. For more than three decades, chemotherapy and radiotherapy have been the only treatments available, but since the discovery of Rituximab, a new era of lymphoma therapy was inaugurated, once it was the first monoclonal antibody approved against the CD20 receptor. The CD20 receptor is an antigen expressed on the B-cells surface, and it is present nearly in all of its maturational development, being absent only in the stages of the pro-B lymphocyte and plasma cells. Furthermore, the CD20 receptor is also present on >90% of the B-cell NHL. These characteristics make this receptor an ideal target for immunotherapy. Besides the success of Rituximab, various mechanisms of resistance have been developed by the tumoral cells against this antibody, such as the decrease of the CD20 expression on B-cells, immunogenicity, structural changes affecting the binding region of the CD20 antibody or alterations in the cell membrane. New anti-CD20 antibodies have been developed to overcome the disadvantages of Rituximab, such as Ofatumumab and Obinutuzumab, presenting a different immunogenicity and binding to the different epitopes on the target with a different affinity. Apart from the progress in the lymphoma therapy, a significant population of patients still succumbs to this disease, as tumoral cells are always changing, making the search for better antibodies a never-ending process. Thus, the aim of this project consisted in the development and characterization of a new antibody against the CD20 receptor. To achieve this goal, the potential of rabbit derived single-domain antibodies (sdAbs) as therapeutic molecules were explored. For that, one rabbit was immunized with the CD20 receptor and an immune VH and VL sdAb library was generated. Then, a subtractive cell phage display screening was used for antibody selection. This approach allowed a specific selection of one sdAb in the VL format against the CD20 receptor in cells. In summary, the strategy explored in the present project resulted in an antibody that, according to its characteristics, could be a promising candidate in the treatment of NHL and other B-cell malignancies.
Os linfomas são a terceira neoplasia mais comum no mundo, sendo os linfomas não-Hodgkin os mais frequentes. Dentro dos linfomas não-Hodgkin, cerca de 85-90% são de células B. Os linfomas desenvolvem-se maioritariamente a partir dos nódulos linfáticos, no entanto podem formar-se a partir de qualquer outro tecido. Durante mais de três décadas, os únicos tratamentos disponíveis para esta doença eram a quimioterapia e a radioterapia. No entanto, com a descoberta do Rituximab, um anticorpo específico para o recetor CD20 presente nos linfomas de células B, começou uma nova era no campo das imunoterapias. O anticorpo Rituximab, uma vez que foi o primeiro a ser descoberto, faz parte da primeira geração de anticorpos anti-CD20. Este é um anticorpo quimérico, uma vez que é constituído por um IgG1 glicosilado composto por uma região constante kappa humana e regiões variáveis leves e pesadas murinas. Este anticorpo atua essencialmente através de três principais mecanismos de ação: citotoxicidade mediada por anticorpos (ADCC); citotoxicidade dependente do complemento (CDC) e indução da apoptose. O recetor CD20 é uma molécula expressa na superfície das células B e está presente na maioria das fases de maturação destas células, com exceção da fase de linfócitos pró-B e nas células plasmáticas. Para além disso, está presente em mais de 90% dos linfomas não-Hodgkin de células B. No entanto, apesar da sua função ainda não ser totalmente conhecida, pensa-se que possa estar envolvido no fluxo de cálcio. Este recetor é uma fosfoproteína não glicosilada com 4 domínios membranares: os domínios N e C terminal que são intracitoplasmáticos e 2 loops adicionais que são extracelulares. Devido a estas suas características, o recetor CD20 é considerado um bom alvo para imunoterapias, nomeadamente terapias para linfomas não-Hodgkin de células B. Apesar do sucesso do Rituximab, têm vindo a ser descritos vários mecanismos de resistência das células tumorais contra este anticorpo, nomeadamente a diminuição da expressão de CD20, mudanças estruturais que afetam o local de ligação do anticorpo à molécula ou alterações na membrana da célula, o que diminuiu a sua eficácia. Este aspeto fez com que fosse necessária a contínua procura por novos anticorpos para colmatar as lacunas do Rituximab. Assim, ao longo dos anos, foram descobertos novos anticorpos específicos para CD20, como o Ofatumumab e o Obinutuzumab, que têm uma diferente imunogenicidade e reconhecem diferentes epítopos na molécula CD20 aos quais se ligam com diferentes afinidades, atuando assim com diferentes mecanismos de ação. Ainda assim, este tipo de doença continua a afetar imensas pessoas, muitas vezes não existindo um tratamento eficaz devido às múltiplas resistências das células tumorais. Tendo em conta este contexto e uma vez que os anticorpos disponíveis no mercado não conseguem resolver estes problemas, a procura por novos anticorpos mais vantajosos continua. Atendendo a esta necessidade, o objetivo deste projeto é o desenvolvimento e caracterização de um anticorpo de pequeno domínio específico para CD20. Os anticorpos de pequenos domínios têm várias vantagens em relação aos IgGs completos, principalmente devido ao seu reduzido tamanho, que faz com que estes anticorpos tenham melhor acesso ao alvo na superfície da célula, para além disso podem ainda ser facilmente expressos em bactérias como proteína. Têm ainda a vantagem, como moléculas terapêuticas, de serem mais estáveis em circulação que os anticorpos completos. Estas características permitem uma administração de maiores quantidades por grama de produto, o que leva a um aumento significativo da potência por dose e na redução do custo de produção. Para o desenvolvimento do projeto, inicialmente foi imunizado um coelho com o péptido CD20 e com células HEK 293T transfetadas com o recetor CD20. Antes de cada imunização, o soro era recolhido para ser avaliado através de ELISA. Ao dia 89, quando foi atingido um elevado título de anticorpos, procedeu-se à recolha do soro final e à eutanásia do coelho. Os ensaios de ELISA permitiram confirmar que o soro extraído após as imunizações estava a reconhecer não só as células Raji, que contêm à sua superfície CD20, mas também o péptido CD20, por outro lado o soro recolhido antes das imunizações não reconhecia nem as células Raji, nem o péptido. Após a eutanásia, foram recolhidos o baço e a medula que são os órgãos onde se encontra um maior número de células plasmáticas que produzem anticorpos. A partir destes órgãos, procedeu-se à extração de ARN usando o reagente Tri. A partir do ARN extraído foi possível sintetizar a primeira cadeia de ADN. O cADN sintetizado foi utilizado para a construção de bibliotecas imunes de pequenos domínios de anticorpos específicos para o recetor CD20, através da amplificação por PCR das famílias de cadeias variáveis leves e pesadas. Esses produtos resultantes da amplificação foram clonados no vetor pComb3x, que é necessário na técnica de phage display, resultando numa biblioteca imune com uma grande diversidade, ideal para este tipo de seleção. Em seguida, as bibliotecas resultantes foram selecionadas através da tecnologia de phage display que permite a seleção de anticorpos específicos para CD20 em células que contenham esse recetor. Esta tecnologia baseia-se na engenharia genética de bacteriófagos e em várias rondas de seleção contra o antigénio e propagação dos fagos. Resumidamente, os fagos vão conter à sua superfície o fenótipo correspondente ao genótipo encapsulado no seu interior que contem a sequência correspondente aos fragmentos de ADN obtidos através da construção das bibliotecas de anticorpos. Seguem-se várias rondas de seleção onde vão sendo eliminados os anticorpos-fagos que não se ligam ou que possuem uma fraca ligação às células que contêm o antigénio, neste caso o recetor CD20, através de várias lavagens. Estas rondas vão sendo repetidas até obtermos uma população de anticorpos específica para o alvo. Para este phage display, foram utilizados 3 tipos de células: células Raji, que está descrito que expressam o CD20, pois são células de linfoma B; células HEK 293T que não expressam CD20 e por isso foram utilizadas para eliminar os anticorpos não específicos para este recetor; células Jurkat, que tal como as anteriores não possuem CD20, pois tratam-se de células T de leucemia aguda e que por isso foram também utilizadas para eliminar anticorpos não específicos. Esta seleção resultou num conjunto de anticorpos-fagos específicos para o recetor CD20, resultado que foi confirmado através de Western Blot. Após a seleção por phage display, foi necessário fazer uma seleção em larga escala para avaliar os anticorpos que melhor eram expressos e que melhor se ligavam ao alvo. Para esta avaliação da expressão foi necessário clonar o ADN resultante dos fagos selecionados por phage display, num outro vetor que permitisse a expressão de proteína, neste caso o vetor pT7-PL. Esta avaliação da ligação e expressão foi feita através de ensaios de ELISA com extrato proteico de células Raji e células Raji. Para além disso, após várias seleções os melhores 10 clones foram sequenciados para avaliar a homologia entre si e verificar os CDRs. Através do alinhamento da sequência de aminoácidos obtida a partir da sequenciação, verificou-se que 9 dos clones eram iguais, restando assim apenas 2 clones. A especificidade dos dois clones selecionados para o recetor CD20 foi avaliada através de Western Blot, sendo que um deles (anticorpo no formato VL) era específico para o recetor pretendido. Estas metodologias levaram à seleção de um anticorpo de pequeno domínio específico para CD20, que devido às suas características únicas e diferentes dos que já existem no mercado poderá ser um promissor candidato para ser usado como agente terapêutico para linfomas de células B que expressem o recetor CD20.