Delineating outcomes of patients with diffuse large b cell lymphoma using the national comprehensive cancer network-international prognostic index and positron emission tomography-defined remission status; a population-based analysis.

The recently devised National Comprehensive Cancer Network International Prognostic Index (NCCN-IPI) appears superior to the revised IPI (R-IPI) in delineating outcome in diffuse large B-cell lymphoma. We examined the outcome of a population-based cohort of 223 consecutive patients treated with R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisolone) or R-CHOP-like immuno-chemotherapy between January 2005 and December 2011 by both the NCCN-IPI and R-IPI, and further stratified outcome by the achievement of both computerized tomography (CT) and positron emission tomography (PET)-CT complete remission (CR), with the latter reassessed using blinded central review by an independent nuclear medicine and radiology specialist. The NCCN-IPI was superior to the R-IPI in identifying patients at very high risk of systemic and/or central nervous system relapse. Notably, both the NCCN-IPI and the R-IPI remained strongly predictive of relapse irrespective of CT or PET-defined remission status following R-CHOP. Patients with high-risk NCCN-IPI scores (≥6) have a dismal outcome following R-CHOP therapy regardless of PET-defined response to R-CHOP. Moreover, such patients appear refractory to salvage chemotherapy and thus require alternative therapeutic approaches, although age and performance status may, for many patients, preclude the safe delivery of a primary intensified regimen. By contrast, patients with NCCN-IPI 1-5 who achieve PET-CR following R-CHOP have excellent outcomes and may merit reduced follow up frequency.

A requirement for calcium in the caspase-independent killing of Burkitt lymphoma cell lines by Rituximab.

The therapeutic monoclonal antibody rituximab has previously been shown to kill B cells in a caspase-independent manner. The signalling pathways underpinning this novel death pathway are unknown. The present study showed that rituximab treatment of Burkitt lymphoma cell lines induced a slow rise in intracellular calcium ([Ca(2+)](i)). This rise was only witnessed in cell lines that were killed by antibody, suggesting a critical role for Ca(2+) in mediating rituximab-driven caspase-independent cell death. Inhibition of the two main intracellular store-located Ca(2+) channels, i.e. the ryanodine and inositol-1,4,5-triphosphate receptor channels by dantrolene and xestospongen-c respectively did not prevent the rise in Ca(2+) seen with rituximab or protect cells from subsequent death. In sharp contrast, inhibition of Ca(2+) entry via plasma membrane channels with (2-aminoethoxy) diphenylborate or SKF-96365 or complete chelation of extracellular Ca(2+) with ethyleneglycol bis (aminoethylether) tetra-acetate inhibited the rise in [Ca(2+)](i) and increased cell viability. Together, these data suggest that ligation of the CD20 receptor with rituximab allows a slow sustained influx of Ca(2+) from the external environment that under certain conditions can lead to cell death.

Outcome of BEAM-autologous and BEAM-alemtuzumab allogeneic transplantation in relapsed advanced stage follicular lymphoma.

The role of haematopoietic stem cell transplantation (HSCT) in relapsed follicular lymphoma remains controversial. This study analysed 126 patients with relapsed, advanced stage follicular lymphoma who received BEAM (BCNU [carmustine], cytarabine, etoposide, melphalan)-alemtuzumab allogeneic HSCT (BEAM-allo) (n = 44) or BEAM-autologous HSCT (BEAM-auto) (n = 82). The BEAM-allo group had a younger median age (48 years vs. 56 years, P < 0.001) but received a higher median number of therapies pretransplant (P = 0.015) compared with the BEAM-auto group. There was a higher non-relapse mortality (NRM) in the BEAM-allo group compared with the BEAM-auto group at 1 year (20% vs. 2%, P = 0.001). Older age and heavily pretreated patients were associated with a higher NRM and poorer survival in the BEAM-allo group. There was, however, a significantly lower relapse rate (20% vs. 43%, P = 0.01) at 3 years with BEAM-alemtuzumab, with no relapses after 2 years, compared with a continued pattern of relapse in the autologous group. No difference in overall survival (OS) (P = 0.99) or disease-free survival (DFS) (P = 0.90) was identified at 3 years, whereas a plateau in OS and DFS with crossing of the survival curves in favour of BEAM-allo group was observed. Furthermore, the ability to re-induce remissions with donor leucocytes provides additional benefit in favour of allogeneic HSCT.

A paraneoplastic manifestation of metastatic breast cancer responding to endocrine therapy: a case report.

BACKGROUND: Many cancers are known to be associated with paraneoplastic syndromes. These syndromes are usually treated by chemotherapy with or without immunosupression but they often respond poorly. There are no published reviews on response to endocrine treatment. CASE PRESENTATION: We report a case of a patient presenting with papillitis, myositis and sensory peripheral neuropathy 18 months before a diagnosis of metastatic oestrogen receptor positive breast cancer was confirmed. The patient was treated with anastrozole which led not only to a decrease of her tumour burden but also to an improvement in her biochemical markers and amelioration of her clinical symptoms. CONCLUSION: This case is an example of breast cancer presenting with paraneoplastic manifestations. It took several months to establish the cause of symptoms in this patient thus illustrating the need for physicians to maintain a high index of suspicion for paraneoplastic syndromes in women presenting with unusual neurological symptoms with no obvious cause.It is a unique case as it illustrates how treatment with an aromatase inhibitor leading to cancer regression can result in an improvement in the paraneoplastic symptoms.

Single-center Series of Bone Marrow Biopsy-Defined Large Granular Lymphocyte Leukemia: High Rates of Sustained Response to Oral Methotrexate.

INTRODUCTION: Large granular lymphocyte (LGL) leukemia is a rare chronic lymphoproliferative disorder, with few large series reported to date. Series using stringent diagnostic criteria incorporating bone marrow biopsy (BMB), immunophenotyping, and T-cell receptor rearrangements are even scarcer. PATIENTS AND METHODS: The present study was a single-center series of 39 patients with LGL leukemia diagnosed using immunohistochemical analysis of BMB samples and flow cytometric and molecular data. RESULTS: With a median follow-up of 3.2 years (range, 1.0-15.1 years), 15 patients (38%) never required treatment. Of the remaining 24 patients requiring treatment, 13 were initially treated with prednisolone, for an overall response rate (ORR) of 84.6% and a median duration of response (DOR) of 13.5 months (range, 5.7-70.3 months). Of the 24 patients, 9 received oral low-dose weekly methotrexate as first-line therapy, with 8 (89%) achieving a hematologic response and a median DOR of 132.7 months (range, 6.7-180.5 months). Another 5 patients received methotrexate after prednisolone failure; all 5 responded, with a median DOR of 14 months (range, 4-96 months). Only 2 patients developed progression during methotrexate therapy, and 4 patients experienced responses lasting ≥ 5 years. CONCLUSION: Single-agent oral methotrexate appears to be highly efficacious, resulting in long response durations and minimal toxicity.

The t(14;18) is associated with germinal center-derived diffuse large B-cell lymphoma and is a strong predictor of outcome.

The t(14;18) is present in a significant proportion of diffuse large B-cell lymphomas (DLBCLs), however, the prognostic effect of the translocation and the relationship with transformed follicular lymphoma remains controversial. To clarify these uncertainties, interphase fluorescence in situ hybridization (FISH) was used to determine the incidence of the t(14;18) in nodal DLBCL, and this was correlated with BCL2 expression, germinal center (GC) immunophenotype, and patient outcome. FISH was performed on paraffin-extracted nuclei from 137 de novo nodal DLBCLs. Eighteen of 137 de novo DLBCLs were t(14;18) positive. The t(14;18) was most commonly associated with the subset of DLBCLs that expressed a GC phenotype, defined as CD10+, BCL6+ (GC-type DLBCL): positive in 14 of 47 (30%) cases, compared with 4 of 89 (5%) in the non-GC group (Pearson's chi(2) = 28.4; P < 0.0001). All cases with a translocation expressed BCL2 protein, however, 40 expressed BCL2 protein without a t(14;18). GC-type DLBCL patients with a t(14;18) had a significantly worse survival compared with GC-type DLBCL patients without the translocation (2-year survivals were 29 and 63%, respectively; P = 0.006). Of the cases without the translocation, BCL2 protein expression did not affect survival. In contrast, in the non-GC group of DLBCLs, BCL2 protein expression reduced the 2-year overall survival from 64% in the BCL2-negative group to 38%, with a median survival of 15.0 months (P = 0.02). In conclusion, the t(14;18) is common in DLBCLs, particularly in GC-type DLBCLs, where the presence of the translocation has a poor prognostic effect. BCL2 protein expression defines a group of non-GC DLBCL patients with a poor prognosis.

Bone marrow trephine biopsy involvement by lymphoma: review of histopathological features in 511 specimens and correlation with diagnostic biopsy, aspirate and peripheral blood findings.

AIMS: This study aimed to evaluate the key features of bone marrow trephine (BMT) biopsy involvement by lymphoma. METHODS: 511 cases were assessed for percentage of marrow involvement, pattern of involvement (diffuse, nodular, paratrabecular, interstitial or intrasinusoidal), presence/absence of granulomas, stromal fibrosis and necrosis, presence/absence of neoplastic/reactive follicles and discordance with other biopsy sites. Correlation with aspirate and peripheral blood findings was made in a subset of 345 patients (167 aspirates, 178 blood). RESULTS: The most frequent subtype was follicular lymphoma (26.2%) followed by extranodal marginal zone (23.1%), lymphoplasmacytic (19.2%), diffuse large B cell (DLBCL) (12.5%), Hodgkin (HL) (5.7%) and mantle cell lymphomas (4.3%). The predominant pattern in follicular lymphoma was paratrabecular. Marginal zone lymphomas of all types and lymphoplasmacytic lymphoma showed a relatively even distribution between diffuse, interstitial, paratrabecular and nodular patterns. The majority of mantle cell lymphoma cases showed either diffuse or nodular patterns. A diffuse pattern was common in DLBCL and Burkitt lymphomas. An intrasinusoidal pattern was seen only in extranodal and splenic marginal zone lymphomas. Granulomas and fibrosis were uncommon in small cell B cell lymphomas but frequent in DLBCL and HL. Aspirate and trephine results concurred in 73.8% of cases overall, but this varied widely between subtypes. Peripheral blood involvement rates by lymphoma also varied, with a mean of 37.1%. CONCLUSIONS: Different lymphomas often demonstrate reliably characteristic architectural patterns of marrow involvement which can help differentiate them even when cytological features do not permit this, and marrow stromal and other background changes may also be useful pointers towards a particular lymphoma subtype.

Hydroxyurea potentiates the caspase-independent killing of B-cell lines by rituximab and GA101.

Anti-CD20 monoclonal antibodies have revolutionized the treatment of non-Hodgkin's lymphoma over the last decade. Unfortunately, a significant number of patients treated by these antibodies exhibit innate or acquired antibody resistance and fail to respond to treatment. Strategies to improve antibody function and overcome resistance include the development of new "engineered" antibodies and the use of new drug combination therapies. In this report, we show that the antimetabolite hydroxyurea significantly enhances the ability of two therapeutic monoclonal antibodies to directly kill some human B-cells. The two anti-CD20 antibodies studied were a clinically well-established type 1 therapeutic antibody, namely rituximab and GA101, an antibody representing the new breed of type 2 glycoengineered monoclonals. Hydroxyurea specifically enhanced the direct caspase-independent killing pathway of both of these antibodies as exemplified by the resistance to broad spectrum caspase inhibitors, lack of internucleosomal DNA laddering, and lack of activation of caspases 3, 8, and 9. Both rituximab and GA101 appear to preferentially kill cells in the G0/G1 cell cycle phase. One of the many reported effects of hydroxyurea is cell arrest in this phase. Arresting antibody-sensitive cells in this stage of the cell cycle by means other than hydroxurea also sensitized the cells to caspase-independent antibody-mediated death, suggesting that the potentiating effect of hydroxyurea may be mediated via its effects upon the cell cycle. The possible combination of hydroxyurea and anti-CD20 monoclonal antibodies may offer new possibilities for combination therapies in the clinic.

Internalisation of uncross-linked rituximab is not essential for the induction of caspase-independent killing in Burkitt lymphoma cell lines.

Characterising the mechanisms underpinning caspase-independent programmed cell death (CI-PCD) induction by uncross-linked rituximab in B-cells may positively impact upon the treatment of disease states in which the classical apoptotic pathway is disabled. The necessity of rituximab internalisation for CI-PCD induction was investigated by flow cytometry and confocal microscopy in human BL cell lines with (e.g. Mutu I) and without (Mutu III) susceptibility to rituximab-induced killing. Flow cytometry demonstrated small, significant and similar amounts of rituximab internalisation by Mutu I cells after 1, 2, 4 and 24 h (p < 0.03, n = 5) and Mutu III cells after 0.5, 2, 4 and 24 h (p < 0.05, n = 4). Confocal microscopy confirmed this. Cytochalasin B and latrunculin A significantly inhibited rituximab-induced CI-PCD (p < or = 0.04, n = 6 and p = 0.01, n = 6, respectively) and internalisation (p = 0.02, n = 5 and p = 0.0002, n = 6, respectively) in Mutu I cells, but confocal microscopy showed no correlation between internalised rituximab and phosphatidylserine exposure. We conclude that rituximab internalisation is not essential for CI-PCD induction in BL cell lines.

Combination chemotherapy followed by autologous stem cell transplant for enteropathy-associated T cell lymphoma.

Enteropathy-associated T cell lymphoma (EATL) is a rare entity associated with coeliac disease, with a poor prognosis due to perforation and gastro-intestinal bleeding during treatment, and a high relapse risk. Six patients were treated with two cycles of IVE (ifosphamide, etoposide, epirubicin), followed by two cycles of high-dose methotrexate (3 g/m(2)) with folinic acid rescue and a BEAM (carmustine, etoposide, cytarabine, melphalan) autograft. Enteral feeding was given throughout treatment. Four patients remain alive in complete remission at 1.83-4.32 years; two have relapsed. Given the historically poor outcome in these patients, this regimen appears very promising in the treatment of EATL.

Ifosphamide, etoposide and epirubicin is an effective combined salvage and peripheral blood stem cell mobilisation regimen for transplant-eligible patients with non-Hodgkin lymphoma and Hodgkin disease.

A total of 143 patients with relapsed (n = 90), primary refractory (n = 32) and first line chemotherapy responsive (n = 21) non-Hodgkin lymphoma (NHL) and Hodgkin disease (HD) were treated with IVE (ifosphamide, etoposide and epirubicin) chemotherapy with the intent to proceed to high-dose therapy with either autologous or allogeneic transplantation, following peripheral blood stem cell mobilisation. A major response (complete/partial response) to IVE was seen in 115 patients (80.4%) with 5-year overall survival (OS) and event free survival (EFS) of 53% and 43%, respectively. Subgroup analysis showed overall response rates of 93.1% for HD with a 5-year OS and EFS of 62% and 52% respectively, while NHL showed response rates of 78.0% with 5-year OS and EFS of 50% and 39% respectively. The median number of CD34 +ve cells mobilised following IVE was 7.86 x 10(6) (range 1.72-42.91 x 10(6)), with 60% mobilising >2 x 10(6)/kg in a single collection. Grade IV neutropenia was seen in 79.6% patients and 77/270 cycles required intravenous antibiotic treatment. We conclude that IVE has a high response rate across a range of refractory and relapsed lymphoma with acceptable toxicity and excellent PBSC mobilising characteristics.

CD95 (Fas) expression is regulated by sequestration in the Golgi complex in B-cell lymphoma.

The CD95 (Fas) molecule transmits apoptotic signals important in B-cell development and the genesis of B-cell lymphoma. We have investigated the surface and intracellular expression of CD95 in Burkitt's lymphoma (BL) cells, an important non-Hodgkin's lymphoma of B-cell origin. Group I BL cells did not express CD95 at the cell surface, but contained high levels of this receptor in the cytoplasm. In contrast, group III BL cells expressed CD95 intracellularly and at the cell surface. In group I and group III BL cells, cytoplasmic CD95 was localized to the Golgi complex, as assessed by confocal immunofluorescence microscopy and subcellular fractionation followed by immunoblotting. Trafficking through the Golgi complex is regulated by elements within the target protein and cellular sorting mechanisms. CD95 contains candidate signals for interaction with trafficking machinery. Group I BL cells can be induced to upregulate surface expression of CD95 following CD40 ligation and certain group I BL cell lines drift invitro to a group III phenotype, with consequent surface expression of CD95. Taken together, these observations show that CD95 can either be retained in the Golgi complex or exported to the cell surface, and suggest that membrane trafficking has an important and previously unrecognized role in regulating CD95 expression in B lymphocytes.

Germinal center phenotype and bcl-2 expression combined with the International Prognostic Index improves patient risk stratification in diffuse large B-cell lymphoma.

The International Prognostic Index (IPI) identifies poor- and good-risk patients with diffuse large B cell lymphoma (DLBCL); however, the majority of patients have an intermediate IPI, with an uncertain prognosis. To determine whether cellular factors can be combined with the IPI to more accurately predict outcome, we have analyzed 177 presentation nodal DLBCLs for the expression of bcl-2 and a germinal center (GC) phenotype (defined by expression of bcl-6 and CD10). P53 gene band shifts were detected using single-stranded conformational polymorphism polymerase chain reaction analysis of exons 5-9 and were correlated with protein expression. In a Cox regression analysis, IPI (R = 0.22, P <.0001) and bcl-2 (R = 0.14, P =.0001) were independent poor prognostic factors and a GC phenotype predicted a favorable outcome (R = -0.025, P =.02). Neither p53 expression nor band shifts had a significant effect on survival. Using the IPI alone, 8% of patients were identified as high risk. Expression of bcl-2 in the intermediate IPI group identified a further 28% of patients with an overall survival comparable to the high IPI group. In the intermediate IPI, bcl-2(-) group, the presence of a GC phenotype improved overall survival to levels approaching the IPI low group. Following this analysis only 15% of patients failed to be assigned to a favorable- or poor-risk group. Sequential addition of bcl-2 expression and GC phenotype into the IPI significantly improves risk stratification in DLBCL. For the 36% of high-risk patients with a 2-year overall survival of 19%, alternative treatment strategies should be considered in future trials.

Rearrangement of the BCL6 locus at 3q27 is an independent poor prognostic factor in nodal diffuse large B-cell lymphoma.

Diffuse large B-cell lymphomas (DLBCL) are a heterogeneous group of tumours, varying in clinical features, immunophenotype and cytogenetics. The aim of this study was to investigate the prognostic significance of BCL6 gene rearrangement at the 3q27 locus in patients with primary nodal disease, and to examine interrelationships with immunophenotype and International Prognostic Index (IPI). We have developed a fluorescent in situ hybridization (FISH)-based technique for the retrospective analysis of the effect of BCL6 gene rearrangements on survival, using nuclei extracted from paraffin-embedded tissue. FISH results were obtained in 111 presentation cases of nodal DLBCL. The IPI was calculated and each case was stained immunocytochemically for BCL6, BCL2 and CD10. 3q27 rearrangements were detected in 25% of cases. BCL2 protein and a germinal centre (GC) phenotype (defined as CD10+, BCL6+) were expressed in 56% and 41% of cases respectively. In multivariate analysis, rearrangement of 3q27 and BCL2 expression and the absence of a GC phenotype were associated with a poor prognosis. These factors can be used in conjunction with the IPI to improve risk stratification in nodal DLBCL.

BEAM-alemtuzumab reduced-intensity allogeneic stem cell transplantation for lymphoproliferative diseases: GVHD, toxicity, and survival in 65 patients.

We report the outcomes of reduced-intensity allogeneic stem cell transplantation using BEAM-alemtuzumab conditioning (carmustine, etoposide, cytosine arabinoside, melphalan, and alemtuzumab 10 mg/d on days -5 to -1) in 6 United Kingdom transplant centers. Sixty-five patients with lymphoproliferative diseases underwent sibling (n = 57) or matched unrelated donor (n = 8) transplantation. Sustained donor engraftment occurred in 60 (97%) of 62 patients. Of the 56 patients undergoing chimerism studies, 35 (63%) had full donor chimerism. Overall, 73% were in complete remission (CR) after transplantation. At a median follow-up of 1.4 years (range, 0.1-5.6 years), 37 remain alive and in CR. Acute graft-versus-host disease (GVHD) occurred in 11 (17%) of 64, grades I-II only. Estimated 1-year transplantation-related mortality (TRM) was 8% for patients undergoing first transplantation but was significantly worse for those who had previously undergone autologous transplantation. Six patients relapsed (estimated 2-year relapse risk, 20%). Histologic diagnosis (mantle cell lymphoma and high-grade non-Hodgkin lymphoma) and age at transplantation (> 46 years) were significantly associated with higher relapse risk and worse event-free survival. Relapse did not occur in any patient who developed acute or chronic GVHD. This study demonstrates that reduced-intensity allogeneic stem cell transplantation for lymphoproliferative diseases using a BEAM-alemtuzumab preparative regimen is associated with sustained donor engraftment, a high response rate, minimal toxicity, and a low incidence of GVHD.