Purpose: The five point Deauville (D) scale is widely used to assess interim PET metabolic response to chemotherapy in Hodgkin lymphoma (HL) patients. An International Validation Study reported good concordance among reviewers in ABVD treated advanced stage HL patients for the binary discrimination between score D1,2,3 and score D4,5. Inter-reader reliability of the whole scale is not well characterised. Methods: Five international expert readers scored 100 interim PET/CT scans from paediatric HL patients. Scans were acquired in 51 European hospitals after two courses of OEPA chemotherapy (according to the EuroNet-PHL-C1 study). Images were interpreted in direct comparison with staging PET/CTs. Results: The probability that two random readers concord on the five point D score of a random case is only 42% (global kappa = 0.24). Aggregating to a three point scale D1,2 vs. D3 vs. D4,5 improves concordance to 60% (kappa = 0.34). Concordance if one of two readers assigns a given score is 70% for score D1,2 only 36% for score D3 and 64% for D4,5. Concordance for the binary decisions D1,2 vs. D3,4,5 is 67% and 86% for D1,2,3 vs D4,5 (kappa = 0.36 resp. 0.56). If one reader assigns D1,2,3 concordance probability is 92%, but only 64% if D4,5 is called. Discrepancies occur mainly in mediastinum, neck and skeleton. Conclusion: Inter-reader reliability of the five point D-scale is poor in this interobserver analysis of paediatric patients who underwent OEPA. Inter-reader variability is maximal in cases assigned to D2 or D3. The binary distinction D1,2,3 versus D4,5 is the most reliable criterion for clinical decision making.

Diffuse large B-cell lymphoma (DLBCL) is the most common form of non-Hodgkin lymphoma (NHL). The current standard of care is the combination of rituximab with cyclophosphamide, doxorubicin, vincristine and prednisone (R-CHOP), but this only results in a 60% over-all 5-year survival rate, thus highlighting a need for new therapeutic approaches. Histone deacetylase inhibitors (HDACi) are novel therapeutics that is being clinically evaluated for combination therapy. Rational selection of companion therapeutics for HDACi is difficult due to their poorly understood, cell-type specific mechanisms of action. To understand these mechanisms better, we developed a pre-clinical model system of response to the HDACi belinostat. Using this model system, we identified two major responses. Resistance, consisting of a reversible G1 cell cycle arrest with little induction of apoptosis; or sensitivity, consisting of mitotic arrest and high levels of apoptosis. In this dissertation, we determine that the induction of G1 cell cycle arrest is due to the increased expression of cyclin dependent kinase inhibitors (CDKi) that bind to and inhibit the cyclin E/CDK2 complex thereby blocking the final repressive phosphorylation steps of Rb protein. Repression of transcriptional elongation blocked CDKi upregulation and prevented G1 cell cycle arrest in belinostat-resistant cells. Additionally, we identified that belinostat arrests sensitive cells prior to metaphase and belinostat-resistant cells slow-down in mitosis but complete the process prior to arresting in G1. The combination of belinostat with the microtubule-targeting agent, vincristine resulted in strong synergistic induction of apoptosis by targeting mitotic progression. Furthermore, this combination prevents polyploidy, a key mechanism of resistance to microtubule targeting agents. Finally, we utilized selective class one HDAC inhibitors to identify the individual contributions of HDACs in the eliciting the responses observed with belinostat treatment. HDAC1&2 inhibition recapitulated the belinostat-resistant phenotype of G1 cell cycle arrest with little apoptosis, in both belinostat-resistant and sensitive cell lines. HDAC3 inhibition resulted in the induction of DNA damage, increased S phase and the induction of apoptosis in belinostat-resistant cells. Belinostat-resistant cells did not have observable effects to HDAC3 inhibitor alone but when combined with vincristine had significantly increased G2/M population at early time points. This suggests that HDAC3 maintains roles in DNA replication and also in mitotic progression. HDAC3 inhibition combined with vincristine resulted in a significant increase in polyploidy, suggesting that HDAC3 might not regulate the expression of apoptotic regulating factors as belinostat does.

No description available.

Classic Hodgkin Lymphoma (cHL) has an annual incidence of 2.4 cases per 100 000 population in the UK, and is one of the most common malignancies diagnosed in young adults aged 15 to 34. The majority of younger patients have a good long-term outcome with between 80 and 90% disease-specific survival but cHL also affects older adults in whom the prognosis is significantly poorer. The role of tumour-associated macrophages (TAM) in cHL has gained much interest, with several studies reporting an association between high numbers of CD68-positive TAM and poor prognosis. There is also a question over the prognostic significance of Epstein-Barr Virus (EBV) infection which is implicated in up to 50% of cHL cases in developed countries. Published data suggests that EBV positivity in elderly patients may be associated with a poorer outcome, whereas in younger adults may be of prognostic benefit. Differences related to age are of interest particularly as an age-related decline in immunity has been linked with the development of certain subtypes of Non-Hodgkin Lymphoma in older patients. In a retrospective study, two separate cohorts of patients with cHL were examined with the aim of identifying: • Differences in the cellular composition of the tumour microenvironment in cHL which has arisen in young and elderly adult patients; • Differences in the cellular composition of the tumour microenvironment in cHL associated with or without EBV infection; • Factors within the tumour microenvironment which may influence prognosis and may be targeted for novel treatments. One group consisted of patients aged between 15 and 34 years at diagnosis and the other, of those aged 60 or over at presentation. Tissue obtained at the time of diagnosis was examined with regard to a number of factors related to the malignant cells and the surrounding microenvironment, including the number and phenotype of macrophages, the number of plasmacytoid dendritic cells and the number of malignant Hodgkin Reed-Sternberg (HRS) cells and non-malignant ‘background’ cells undergoing apoptosis. Comparisons were made between the two age groups, also taking into account the EBV-status of tumours, cHL subtype and gender. Results confirmed the current understanding that EBV-positive cHL is more common in older patients and has a strong, but not exclusive, association with the MCHL subtype. In addition, a strong link between young males and EBV-positive disease was shown. Macrophages were found to vary between the two age groups, in number and phenotype and there were clear differences associated with the presence or absence of EBV infection. While no definite link with outcome and macrophages was identified it was apparent that the implications of macrophages in the tumour microenvironment may differ between the two age groups. The number of apoptotic cells correlated closely with the number of macrophages and in the young the number of HRS cells was associated with prognosis. Investigation of the tumour microenvironment is complex and caution is needed in interpreting studies which do not differentiate between patients according to age, as tumour characteristics may have variable implications in different age groups. In this thesis a number of clinicopathological differences were identified between the two age groups. These point to the need for further larger studies to delineate how such age-related differences may or may not be associated with immune function and how this information could be translated into treatments to improve outcomes.

Pacientes com linfoma de Hodgkin (LH) tratados com poliquimioterpia com adriamicina, bleomicina, vincristina e doxorrubicina (ABVD) apresentam resposta terapêutica distinta. Para aprimorar a avaliação prognóstica e a abordagem terapêutica em LH objetivamos avaliar o valor prognóstico da PET-FDG após 2 ciclos de ABVD (PET2) em pacientes com LH. Foram incluídos nesse estudo prospectivo 115 pacientes com diagnóstico recente de LH no período de agosto de 2005 a dezembro de 2007. Os pacientes foram estadiados com exame clínico, laboratorial, tomografia computadorizada e PET-FDG (PET0). Todos os pacientes foram tratados com ABVD e aqueles com massa tumoral extensa foram tratados com radioterapia associada. Após dois ciclos de ABVD os pacientes foram submetidos a PET2. Nenhum tratamento foi alterado baseado na PET2. Foi avaliado o valor prognóstico dos fatores clínicos, Índice Prognóstico Internacional (IPI) e PET2 em relação à sobrevida livre de eventos (SLE) em três anos. Dos 104 pacientes que foram avaliados, 82 atingiram remissão completa e 22 pacientes apresentaram falha de tratamento durante a mediana de 36 meses de acompanhamento. A SG e SLE em três anos foi de 94,2% e 74,2% respectivamente. A SLE em três anos da PET2 positiva foi de 54,3%, enquanto da PET2 negativa foi de 90,5% (p< 0.001). Na análise de subgrupos de pacientes com estádio precoce, avançado, IPI baixo e alto risco, a PET2 também apresentou correlação estatisticamente significativa com o prognóstico. Concluímos que a PET2 é o melhor fator prognóstico independente na avaliação de pacientes com LH / Patients with Hodgkin lymphoma (HL) treated with poliquimioteraphy with adriamycin, bleomycin, vincristine and doxorubicin (ABVD) have distinct therapeutic response. In order to improve the prognostic assessment and therapeutic approach in HL we have evaluated the prognostic value of FDG-PET after 2 cycles of ABVD (PET2). Were included in this prospective study 115 patients with newly diagnosed LH in the period of August 2005 to December 2007. The patients were staged with physical examination, laboratory, CT and PET-FDG (PET0). All patients were treated with ABVD and those with extensive tumor were treated with radiotherapy associated. After two cycles of ABVD patients underwent PET2. No treatment was changed based on PET2. We assessed the prognostic value of clinical factors, international prognostic score (IPS) and PET2 in relation to event-free survival (EFS) in three years. Of the 104 patients who finalized the evaluation, 82 achieved complete remission and 22 patients experienced treatment failure during the median of 36 months of follow-up. The EFS at three years was 74.2%. EFS in three years of PET2 positive was 54.3%, while the PET2 negative was 90.5% (p <0.001). In subgroup analysis of patients with early stage, advanced, low and high risk IPS, PET2 also showed significant correlation with the prognosis. We conclude that the PET2 is the best independent prognostic factor in the evaluation of overall patients with LH, or in subgroups of early, advance; low and high risk of HL

No description available.

Pacientes com linfoma de Hodgkin (LH) tratados com poliquimioterpia com adriamicina, bleomicina, vincristina e doxorrubicina (ABVD) apresentam resposta terapêutica distinta. Para aprimorar a avaliação prognóstica e a abordagem terapêutica em LH objetivamos avaliar o valor prognóstico da PET-FDG após 2 ciclos de ABVD (PET2) em pacientes com LH. Foram incluídos nesse estudo prospectivo 115 pacientes com diagnóstico recente de LH no período de agosto de 2005 a dezembro de 2007. Os pacientes foram estadiados com exame clínico, laboratorial, tomografia computadorizada e PET-FDG (PET0). Todos os pacientes foram tratados com ABVD e aqueles com massa tumoral extensa foram tratados com radioterapia associada. Após dois ciclos de ABVD os pacientes foram submetidos a PET2. Nenhum tratamento foi alterado baseado na PET2. Foi avaliado o valor prognóstico dos fatores clínicos, Índice Prognóstico Internacional (IPI) e PET2 em relação à sobrevida livre de eventos (SLE) em três anos. Dos 104 pacientes que foram avaliados, 82 atingiram remissão completa e 22 pacientes apresentaram falha de tratamento durante a mediana de 36 meses de acompanhamento. A SG e SLE em três anos foi de 94,2% e 74,2% respectivamente. A SLE em três anos da PET2 positiva foi de 54,3%, enquanto da PET2 negativa foi de 90,5% (p< 0.001). Na análise de subgrupos de pacientes com estádio precoce, avançado, IPI baixo e alto risco, a PET2 também apresentou correlação estatisticamente significativa com o prognóstico. Concluímos que a PET2 é o melhor fator prognóstico independente na avaliação de pacientes com LH / Patients with Hodgkin lymphoma (HL) treated with poliquimioteraphy with adriamycin, bleomycin, vincristine and doxorubicin (ABVD) have distinct therapeutic response. In order to improve the prognostic assessment and therapeutic approach in HL we have evaluated the prognostic value of FDG-PET after 2 cycles of ABVD (PET2). Were included in this prospective study 115 patients with newly diagnosed LH in the period of August 2005 to December 2007. The patients were staged with physical examination, laboratory, CT and PET-FDG (PET0). All patients were treated with ABVD and those with extensive tumor were treated with radiotherapy associated. After two cycles of ABVD patients underwent PET2. No treatment was changed based on PET2. We assessed the prognostic value of clinical factors, international prognostic score (IPS) and PET2 in relation to event-free survival (EFS) in three years. Of the 104 patients who finalized the evaluation, 82 achieved complete remission and 22 patients experienced treatment failure during the median of 36 months of follow-up. The EFS at three years was 74.2%. EFS in three years of PET2 positive was 54.3%, while the PET2 negative was 90.5% (p <0.001). In subgroup analysis of patients with early stage, advanced, low and high risk IPS, PET2 also showed significant correlation with the prognosis. We conclude that the PET2 is the best independent prognostic factor in the evaluation of overall patients with LH, or in subgroups of early, advance; low and high risk of HL

<p>The relationship between Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) is not entirely elucidated and a clonal relation may be present more often than previously believed. Mechanisms of tumour progression and resistance to therapy are poorly understood.</p><p>Between 1974 and 1994 all individuals in Sweden with both HL and NHL were identified. Thirty-two cases were studied using clinical, histopathological and immunohistochemical methods. The second lymphoma often appeared in an aggressive clinical form and a significant correlation between the expression of p53 and LMP-1 in the first and second lymphoma was demonstrated.</p><p>The treatment outcome for 307 patients with advanced stages of HL, in an unselected population was in accordance with the treatment results of large centres world-wide. Some patients were successfully selected for a shorter chemotherapy-regimen without inferior treatment results.</p><p>In 124 patients with relapse, the survival of those primarily treated with radiotherapy according to the National guidelines was in accordance with the survival of patients of initially advanced stages. A worse outcome was found for those who received both chemotherapy and radiotherapy initially, probably because of a higher frequency of bulky disease in this group. </p><p>Immunohistochemical analysis of the tumour suppressor protein p53 and retinoblastoma protein (Rb) of paired samples at diagnosis and at relapse in 81 patients did not reveal any specific staining pattern affecting survival.</p><p>A novel B-cell line (U-2932) was established from a patient with a diffuse large B-cell lymphoma previously treated for advanced stage and subsequent relapses of HL. An identical rearranged IgH gene was demonstrated in tumour cells from the patient and in U-2932. A p53 point mutation was detected and over-expression of the p53 protein was found. A complex karyotype with high-level amplifications of the chromosomal regions 18q21 and 3q27, i.e. the loci for <i>bcl-2</i> and <i>bcl-6</i> were demonstrated. </p>

The relationship between Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) is not entirely elucidated and a clonal relation may be present more often than previously believed. Mechanisms of tumour progression and resistance to therapy are poorly understood. Between 1974 and 1994 all individuals in Sweden with both HL and NHL were identified. Thirty-two cases were studied using clinical, histopathological and immunohistochemical methods. The second lymphoma often appeared in an aggressive clinical form and a significant correlation between the expression of p53 and LMP-1 in the first and second lymphoma was demonstrated. The treatment outcome for 307 patients with advanced stages of HL, in an unselected population was in accordance with the treatment results of large centres world-wide. Some patients were successfully selected for a shorter chemotherapy-regimen without inferior treatment results. In 124 patients with relapse, the survival of those primarily treated with radiotherapy according to the National guidelines was in accordance with the survival of patients of initially advanced stages. A worse outcome was found for those who received both chemotherapy and radiotherapy initially, probably because of a higher frequency of bulky disease in this group. Immunohistochemical analysis of the tumour suppressor protein p53 and retinoblastoma protein (Rb) of paired samples at diagnosis and at relapse in 81 patients did not reveal any specific staining pattern affecting survival. A novel B-cell line (U-2932) was established from a patient with a diffuse large B-cell lymphoma previously treated for advanced stage and subsequent relapses of HL. An identical rearranged IgH gene was demonstrated in tumour cells from the patient and in U-2932. A p53 point mutation was detected and over-expression of the p53 protein was found. A complex karyotype with high-level amplifications of the chromosomal regions 18q21 and 3q27, i.e. the loci for bcl-2 and bcl-6 were demonstrated.

Made available in DSpace on 2015-07-22T20:50:12Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-08-25 / Os antígenos cancer/testículo (CTAs) são considerados promissores alvos para abordagens de imunoterapia em câncer devido à sua alta imunogenicidade e padrão de expressão praticamente restrito a tecidos tumorais (estão também expressos em células germinativas do testículo, placenta e ovário fetal). Apesar do padrão de expressão dos CTAs estar bem estabelecido em carcinomas, pouco se sabe sobre a expressão desses antígenos em neoplasias linfóides como Linfomas de Hodgkin (LH) e Linfomas não-Hodgkin (LNH). Objetivo: Avaliar o potencial desses antígenos específicos tumorais como candidatos à imunoterapia em linfomas, através da análise de expressão protéica dos CTAs e avaliação da resposta imune humoral espontânea contra esses antígenos, correlacionando os achados com os dados clínicos e prognóstico. Métodos: Três blocos de Tissue Microarray (TMA) foram construídos a partir de 38 amostras teciduais de LH clássico e 106 de LNH obtidos nos arquivos do Departamento de Anatomia Patológia da UNIFESP. As lâminas de TMA foram submetidas a um painel de imunohistoquímica para 9 CTAs, a saber: MAGE-A1, MAGE-A3, MAGE-A4, MAGE-A10, CT7, CT10, NY-ESO-1, LAGE e GAGE. A avaliação da imunidade humoral espontânea foi realizada em 97 amostras de soro de pacientes com LNH ao diagnóstico (59 dos quais foram incluídos na casuística do TMA), utilizando-se a técnica de ELISA em um painel mais amplo de CTAs (MAGEA1, MAGE-A3, MAGE-A4, MAGE-A10, NY-ESO-1, CT7, CT10, CT24, CT45, CT46, CT47, CT63, CT83, SSX-1, SSX-2, SSX-4, LAGE-1, GAGE-2, SAGE-1, XAGE-1). Resultados: De forma global, houve baixa expressão de CTAs nas amostras analisadas, visto que apenas 21,1% das amostras de LH e 11,3% das amostras de LNH apresentaram positividade para pelo menos 1 dos CTAs incluídos no painel de imunohistoquímica. MAGE-A (18,4%) e CT7 (13,2%) foram os CTAs mais frequentemente expressos em LH, enquanto MAGE-A (6,6%), GAGE (5,7%) e NY-ESO-1 (4,8%) foram os mais expressos em LNH. Apesar de não ter sido encontrada diferença estatisticamente significante na expressão de CTAs entre os subgrupos clínicos de LH, houve maior frequência de positividade nos pacientes com estadiamento avançado (28,6%), comparado àqueles com estadiamento inicial (11,8%). Nos subgrupos clínicos de LNH, a frequência de expressão de CTAs foi maior nos linfomas agressivos (14,9%) em relação aos indolentes (3,1%), nos linfomas difusos de grandes células B (LDGCB) (16,1%) comparado aos não-LDGCB (6,0%) e no subgrupo que não atingiu resposta completa (15,0%) comparado àqueles que obtiveram resposta completa (6,8%). Entretanto, as diferenças não foram estatisticamente significantes em nenhum desses subgrupos. Um achado inesperado no presente estudo foi a expressão mais frequente de CTAs no subgrupo de LNH com estadiamento inicial (I e II) comparado ao subgrupo com estadiamento avançado (III e IV). Apesar da diferença encontrada na análise de sobrevida entre o grupo de pacientes que não apresentaram expressão de CTA (sobrevida mediana de 65 meses) e aqueles que apresentaram expressão de pelo menos 1 CTA (sobrevida mediana de 11 meses), a diferença não foi estatisticamente significante (p=0.0947). A resposta humoral espontânea contra pelo menos 1 CTA do painel foi encontrada em 19,6% dos pacientes com LNH. CT47 foi o CTA mais frequentemente expresso (7.2%), seguido do CT45 (5.1%), NY-ESO-1 (5.1%) e MAGE-A4 (5.1%). Os CTAs foram mais frequentemente expressos em LNH de células B (21.2%) comparado aos LNH de células T (6.2%) (p=0.048). Não houve diferença estatisticamente significante na resposta humoral anti-CTAs em qualquer dos outros subgrupos clínicos de LNH. Conclusão: De forma geral, houve baixa expressão protéica de CTAs em nossa casuística de LH e LNH com o painel utilizado. Apesar dos limitados dados disponíveis na literatura, esses achados são concordantes com a maioria dos estudos realizados utilizando RT-PCR e/ou imunohistoquímica. Não houve correlações estatisticamente significantes entre expressão de CTAs e parâmetros clínicos ou prognósticos no presente estudo. A reatividade sérica contra os CTAs testados ocorreu em níveis semelhantes ao da expressão protéica em LNH, sugerindo que os pacientes com LNH são capazes de montar resposta imune humoral específica contra CTAs. / Cancer/testis antigens (CTAs) are expressed in a variety of malignant tumors but in normal adult tissues solely in testicular germ cells. Based on this tumor-associated expression pattern, these antigens are potential targets for immunotherapy. Though carcinomas have been extensively analyzed, less is known about lymphoid malignancies such as lymphomas. Aims: To evaluate the potential of tumor specific antigens as candidates for immunotherapy in lymphomas throughout CTA protein expression and spontaneous humoral immune response analyses. We also aim to investigate clinical correlations and prognostic impact of CTAs expression in lymphomas. Methods: Tissue microarray was generated from 38 Hodgkin´s lymphoma (HL) and 106 non- Hodgkin´s lymphoma (NHL) archival cases. Immunohistochemistry (IHC) was done using a panel of 9 monoclonal antibodies against CTAs. Spontaneous humoral immune response analysis against a larger CTA panel was performed in 97 untreated NHL patient samples, including 59 cases from the TMA cohort, using ELISA technique. Results: We found overall low expression of CTAs in our series of HL (21.1%) and NHL (11.3%) TMAs, being MAGE-A (18.4%) and CT7 (13.2%) the most frequently expressed CTAs in HL, and MAGE-A (6.6%), GAGE (5.7%), NY-ESO-1 (4.8%) and CT7 (4.8%) the most frequently expressed CTAs in NHL. Although we did not find statistically significant difference in CTA expression among the clinicopathological subgroups of HL, CTA positivity was higher in advanced stage (28.6%) compared to early stage patients (11.8%). Among NHL, we found higher CTA expression in aggressive lymphomas (14.9%) compared to indolent lymphomas (3.1%), DLBCL (16.1%) compared to non-DLBCL (6.0%) and in non-complete response group (15.0%) compared to those who achieved complete response (6.8%), but it was not statistically significant. Unexpectedly, early stage disease (19.5%) had higher CTA expression than advanced stage NHL (6.2%). Despite the difference found in survival analysis between NHL patients that presented no CTAs expression (median OS 65 months) and those who expressed at least one CTA (median OS 11 months), it did not reach statistically significant difference (p=0.0947). Serum reactivity against at least 1 CTA was observed in (19.6%) of NHL patients, being more frequent in B-cell lymphomas (21.2%) then T-cell lymphomas (6.2%) (p=0.048). CT47 was the most frequently expressed CTA (7.2%), followed by CT45 (5.1%), NY-ESO-1 (5.1%) and MAGE-A4 (5.1%). Grouping the MAGE-A family similarly to the TMA analysis, we found positivity in 8.2% of NHL serum samples. Among DLBCL, CT45 and NY-ESO-1 were the most frequently expressed CTAs, being positive in 4/50 (8.0%) and 3/50 (6.0%), respectively. Conclusion: We found overall low expression of CTAs in our series of HL and NHL TMAs, and low reactivity against CTA in our serum samples. Our results demonstrated a slightly higher frequency of humoral response against most CTAs included in both TMA and ELISA panel compared to their expression by TMA. Considering that generally a small proportion of patients expressing CTAs develop specific humoral response, it is possible that CTA expression by TMA could be underestimated due to the focal expression pattern in some patients. Therefore, using an extensive panel of antibodies and large TMA and serum cohorts of lymphoma patients, we could not identify CTA candidates for immunotherapy in HL and NHL. / TEDE / BV UNIFESP: Teses e dissertações