[Treatment of diffuse large B-cell lymphoma]. / Behandlung diffus-grosszelliger B-Zell-Lymphome.

Diffuse large B-cell lymphoma represent 40% of all lymphoma. The development of dose-dense chemotherapeutic regimens and the application of the monoclonal CD20 antibody rituximab improve the prognosis significantly. Evaluation of clinical risk factors (age, stage, LDH, ECOG performance status, number of extranodal involvement) at initial diagnosis are the most important approaches for risk stratification that allows risk adapted modifications of the standard R-CHOP regimen.

Report of a workshop on malignant lymphoma: a review of molecular and clinical risk profiling.

Molecular genetic analysis adds important information for lymphoma biology and classification, but the latter is challenged by recent improvement of combined chemo-immunotherapy. In aggressive lymphoma, molecular profiling identifies risk groups with certain genetic background but still the International Prognostic Index (IPI) remains the most important clinically applicable predictor of outcome. In follicular lymphoma (FL), the importance of the meshwork of bystander cells becomes increasingly evident. As combined immuno-chemotherapy improved the prognosis of the patients, several clinical trials indicated that the FLIPI still efficiently discriminates patients at risk for transformation and relapse, although several mechanisms of transformation seem to exist. In mantle cell lymphoma it has been proven that pathogenesis and prognosis mainly depend on deregulation of the cell cycle. A reliable clinical risk score could be established.

Successfully treated Candida krusei infection of the lumbar spine with combined caspofungin/posaconazole therapy.

Candidal vertebral osteomyelitis represents an extremely rare invasive mycosis and can be difficult to treat due to poor drug penetration into bony tissue. We report on a case of vertebral osteomyelitis caused by Candida krusei in a patient who had neutropenia as a result of chemotherapy for acute myelogenous leukaemia. The patient received prophylactic liposomal amphotericin B during chemotherapy but became febrile and experienced severe lumbar pain. Magnetic resonance imaging revealed vertebral osteochondrosis. C. krusei was recovered from blood cultures and voriconazole monotherapy was initiated but proved unsuccessful. The patient was then started on caspofungin monotherapy, which was discontinued after Candida krusei was no longer recoverable from blood cultures. However, as lumbar pain increased and spinal biopsy confirmed the presence of Candida krusei, caspofungin therapy was resumed. Oral posaconazole was added to the regimen when the patient did not improve after 30 days of caspofungin therapy. Combined antimycotic therapy resulted in a successful outcome.

Treatment of CNS dissemination in systemic lymphoma.

The frequency of central nervous system (CNS) dissemination in non-Hodgkin's lymphoma (NHL) varies and is dependent on NHL histology. More than 50% of patients with CNS involvement have advanced and progressive systemic disease. While CNS involvement at initial diagnosis may be treated curatively, treatment of CNS involvement in systemic relapsing or refractory lymphoma is challenging and most often palliative. Due to a paucity of randomized trials, treatment of lymphomatous metastases is not standardized. Nonetheless, treatment of LM entails administration of both CNS-directed and systemic chemotherapy that often includes high-dose chemotherapy regimens with stem cell support.

[Treatment of aggressive lymphomas]. / Therapie grosszelliger Lymphome.

The age-adjusted International Prognostic Index (IPI) allows the definition of clinically relevant subgroups in younger patients (< or = 60 years) with aggressive lymphomas. Six cycles of cyclophosphamide, doxorubicin, vincristine, and prednisone-21 (CHOP-21) with rituximab (R-CHOP-21) achieved an event free survival after 3 years of > 90% in patients with a very good prognosis (IPI 0, without bulk). In contrast, in patients from the less favourable groups (all IPI 1, IPI 0 with bulk only), the same therapy resulted in an event-free survival at 3 years of less than 80%. This requires improvement. Young patients with a worse risk profile have a survival probability of 60% after 5 years. Dose dense therapeutic regimens and autologous stem cell transplantation are being applied to this patient group in clinical trials. For patients > 60 years, the advantage of combined immunochemotherapy has been clearly documented. For patients > 60 years from all IPI groups, the best results have so far been achieved with six courses of R-CHOP-14 followed by two additional applications of rituximab.

Improved outcome of zygomycosis in patients with hematological diseases?

Zygomycosis is an opportunistic fungal infection that is increasingly reported in hematological patients. We describe 2 cases of successfully treated rhino-cerebral zygomycosis and give an overview of 120 patients from the literature with underlying hematological or oncological disorders. These data document the improved survival in sinus (15/17 patients surviving) and cutaneous (6/9 patients surviving) disease. Hematological patients with pulmonary (9/30 patients surviving) or disseminated (4/38 patients surviving) zygomycosis still have a poor prognosis. The clinical course of sinus-orbital involvement (4/11 patients surviving) follows sinus-cerebral (2/3 patients surviving) or cerebral (3/6 patients surviving) disease. Besides deoxycholate amphotericin B (AmB) (24/62 patients surviving), patients seem to benefit from liposomal amphotericin B (L-AmB) (10/16 patients surviving) or sequential AmB/L-AmB treatment (6/8 patients surviving). Alternative treatment options lead only in a few patients to success.

CSF evaluation in primary CNS lymphoma patients by PCR of the CDR III IgH genes.

BACKGROUND: Cytologic evaluation of CSF does not consistently detect malignant cells in patients with primary CNS lymphoma (PCNSL). The potentially more sensitive molecular assessment of monoclonality has not been shown in CSF samples. METHODS: The authors studied nested PCR of the complementary determining region III (CDR III) on 76 CSF specimens of patients with PCNSL. Patients with systemic disseminated B-cell non-Hodgkin's lymphoma (n = 17) and 17 patients with no history of lymphoma were compared. PCR products were evaluated by automated fluorescent fragment analysis (ALF). RESULTS: In 68 patients with PCNSL, the authors analyzed the first obtained CSF sample. Nevertheless, 60 patients were taking corticosteroids. In 16 PCNSL samples, amplifiable DNA was not yielded. Taking into account that at least two independent assays have to be performed, CDR III PCR consistently revealed monoclonal products in eight PCNSL and polyclonal results in 52 PCNSL specimens. CDR III PCR detected no monoclonal PCR products in patients without history of lymphoma. In 10 patients with PCNSL, the PCR result and the CSF cytology were discordant. Concerning therapeutic impact, leptomeningeal tumor spread did not predict tumor response in this group of patients with PCNSL. CONCLUSIONS: This study performed CDR III PCR as a routine diagnostic technique applicable even on CSF samples with low cell counts. These data present low incidence of leptomeningeal involvement in this subset of pretreated PCNSL patients. Because the CSF evaluation did not predict outcome in our patients, further analysis in patients with PCNSL should focus on CSF samples that are obtained very early after diagnosis.

Prospective BCR-ABL analysis by polymerase chain reaction (RT-PCR) in adult acute B-lineage lymphoblastic leukemia: reliability of RT-nested-PCR and comparison to cytogenetic data.

The reliability of routine BCR-ABL RT-nested-PCR was evaluated in 1453 B-lineage ALL or hybrid leukemia at initial diagnosis by RT-nested-PCR. All BCR-ABL-positive (n = 642) and 176 BCR-ABL-negative samples underwent a second RT-PCR. In 518 patients, karyotyping and/or FISH was compared to the BCR-ABL status. The second RT-PCR revealed in 155/642 initially positive samples a divergent result (153 BCR-ABL-negative, two other transcripts) that in most cases turned out to be caused by contaminations in the first RT-nested-PCR. Confirmatory RT-PCR detected 2/176 false negative first RT-nested-PCR results. Thirty-nine specimens remained ambiguous despite different RT-PCR approaches. As far as cytogenetic evaluation and FISH is available (n = 23), the majority but not all patients with an ambiguous RT-PCR result were Ph-negative (n = 18). RT-nested-PCR and cytogenetics yielded in 346 of 383 evaluable samples a concordant result. Differing results are given and account in part to the lower sensitivity of karyotyping. Taken together, confirmed RT-PCR detected BCR-ABL fusion transcripts consistently in 487 out of 1453 ALL samples (c-ALL: 43%, pre-B ALL: 34%, pro-B ALL: 5%, B-ALL: 0%, hybrid leukemia: 5/11). Since false positive initial RT-nested-PCR data were frequent, either confirmatory second RT-PCR or FISH analysis is warranted to guarantee sensitive and reliable results of utmost clinical relevance.

Comparison of ethidium bromide-stained agarose gel electrophoresis and automated fragment analysis for evaluation of IgH gene products.

In acute lymphoblastic leukemia (ALL) patients, automated fluorescence fragment analysis (ALF) has been reported to improve the monoclonality detection rate of immunoglobulin heavy chain genes (IgH) polymerase chain reaction (PCR) analysis. This study performed complementary determining region (CDR) I and III PCR on samples from 135 patients with B-cell neoplasias and 25 healthy controls. The value of ALF was investigated in comparison to the widely used ethidium bromide (ETB)-stained agarose gels (AGGE). ETB-stained AGGE detected monoclonal CDR III PCR products in 53/72 ALL, in 22/34 non-Hodgkin's lymphoma (NHL), 13/22 multiple myeloma (MM), and 2/7 monoclonal gammopathies (MGUS). ALF identified monoclonal CDR III amplificates in 55/72 ALL, 23/34 B-NHL, 14/22 MM, and 2/7 MGUS. AGGE achieved clonal CDR I PCR results in 30/64 samples, while ALF detected 34 clonal CDR I product patterns. Taking together, ETB-stained AGGE revealed monoclonality in 120/199 PCR products versus 129/199 by ALF. Compared with AGGE and ETB-staining, ALF offers a slightly increased sensitivity and can be recommended for the evaluation of difficult samples.

The preparation and execution of saccadic eye and goal-directed hand movements in patients with Parkinson's disease.

The oculomotor and manual motor systems were studied in a two-segment movement task in a group of patients with Parkinson's disease and in age matched normal controls. In order to avoid reflexive motor movements the selection of the correct motor sequence was derived from the interpretation of symbolic (coloured) cues. The latencies and dynamics of eye and hand (pointing) movements performed during the first (fixed) movement segment were measured and the planning and execution processes were manipulated by varying the complexity of the second movement segment relative to the first (with regard to direction and amplitude). The results showed that the eye and hand movements made by patients with Parkinson's disease were not impaired in the initiation of the first movement segment. Interestingly, both Parkinson's patients and controls showed increased eye and hand reaction time latencies for the first movement when the second movement was in the direction opposite to the first. This indicates that the complexity of the second movement influences the execution of the first movement, and importantly that complexity affects motor initiation and execution processes in both normal subjects and in patients with Parkinson's disease. The execution of hand movements was found to be impaired in patients with Parkinson's disease as indicated by a reduced peak velocity of manual pointing responses when compared to age matched controls. By contrast, no differences were found in the dynamics of saccadic eye movements. This dissociation is consistent with the notion that the skeletomotor loop passes through the functionally corresponding portions of the basal ganglia independently of the oculomotor loop. Together, these results demonstrate that Parkinson's patients are able to generate multiple non-reflexive eye and hand movements and that the observed (manual) motor deficits are specific to the processes of motor execution.

Detection of immunoglobulin heavy chain gene rearrangements in hematologic malignancies.

B-cell precursors undergo a unique somatic immunoglobulin heavy chain gene rearrangement process. The generated VHDHJH junction is a successful marker in lymphoproliferative malignancies at initial diagnosis for detection of clonality and during treatment for monitoring minimal residual disease. VHDHJH errors are often involved in recurring structural chromosomal aberrations of lymphoid malignancies, with consequent deregulated expression of the juxtaposed oncogenes, e.g., c-myc, bcl-2 or CCND1. Besides cytogenetics, a variety of molecular techniques are becoming increasingly established, including Southern blotting, PCR and real-time PCR, as well as fluorescence in situ hybridisation. Different approaches may be chosen to evaluate lymphoid malignancies either at diagnosis or follow-up in the light of increasing relevance and proven clinical utility.

Analysis of chimerism during the early period after allogeneic peripheral stem cell transplantation.

As there are few reports on early evaluation of chimerism, we assessed fluorescence short tandem repeats (STR) by polymerase chain reaction (PCR) assays to analyse donor and recipient characteristics at early time points after peripheral stem cell transplantation (PBSCT). Peripheral blood of 13 patients was analysed in 1- to 2-day intervals starting from the day of PBSCT. Donor and recipient allelic patterns were determined by a commercially available multiplex STR assay that simultaneously evaluates four or five gene loci. Mixed chimerism appeared in all patients during days 1-9 after transplantation and preceded haematologic engraftment for 3-12 days. Even patients without myeloablative conditioning therapy (n=4) revealed donor allelic patterns within 1-5 days. Nine patients changed during the following days to a complete donor allelic pattern and had an uncomplicated post-transplant disease course. Four patients did not consistently retain complete donor chimerism; two of them relapsed within the next 3 months, one died from septicemia within 7 days, and the fourth, transplanted for aplastic anaemia, is still in complete remission. Overall, STR analysis using a simple and comparatively cheap multiplex system permits the detection of chimerism very early after transplantation and may provide relevant information that correlates with the clinical follow-up.

Serological and nucleotide sequence analysis of HLA-A*6812.

We describe a novel HLA-A allele, HLA-A*6812. HLA-typing of a patient was performed by serology and by DNA-based analyses. Cloning and sequencing of exons 2 and 3 revealed that the genotype consisted of a common HLA-A*0101 allele and the novel HLA-A*6812 allele. HLA*6812 is identical to HLA-A*68012 except for a single non-synonymous nucleotide substitution leading to an exchange of a threonine to an isoleucine residue at position 142 of the HLA-class I alpha chain. The allele was also found in the patient's mother. The novel HLA-A68 molecule is recognised by some but not all of our HLA-A28-specific sera. These results confirm that position 142 is important for the serological properties of the HLA-A molecule.

Detection of immunoglobulin heavy chain genes rearrangements in B-cell leukemias, lymphomas, multiple myelomas, monoclonal and polyclonal gammopathies.

Polymerase chain reaction (PCR) based assays were found to be a realistic alternative to Southern blot hybridization for the assessment of clonal immunoglobulin heavy chain gene rearrangements. However, a comparison of the different PCR based studies reveals considerable variation in experimental design and marked differences in the reported results. This study compared different single- and double-step PCR assays relying on various FR3, FR2, FR1 and JH based primers for the detection of B cell clonality in acute lymphoblastic leukemias (ALL), non-Hodgkin's-lymphoma (NHL), multiple myeloma (MM), monoclonal gammopathies of unknown significance (MGUS) and three polyclonal gammopathies (PG). The highest monoclonality rate was observed using seminested CDR-III region amplification. This method achieved a monoclonal product in 6 of 13 pro-B ALL 21 of 29 c-ALL, 7 of 8 pre-B-ALL, 18 of 21 B-ALL, 14 of 17 B-NHL (intermediate or high grade) with bone marrow involvement, 0 of 9 B-NHL without bone marrow involvement, 9 of 9 low grade B-NHL (immunocytoma and including chronic lymphocytic leucemia), 13 of 19 MM, 2 of 9 MGUS, and 0 of 3 PG. Additional monoclonality was detected with nested CDR I PCR in 1 pro-B-ALL, 1 c-ALL, and 2 MM. CDR III IgH PCR has been confirmed as an efficient method for determining clonality in B-cell neoplasias. Some additional monoclonal products can be seen with CDR I-based PCR. Detection of monoclonality depends on the maturation grade of the neoplastic B-cell population.

Phase II trial of N-(phosphonacetyl)-L-aspartate (PALA), 5-fluorouracil and recombinant interferon-alpha-2b in patients with advanced gastric carcinoma.

The aspartate transcarbamoylase inhibitor, N-(phosphonacetyl)-L-aspartate (PALA), synergistically enhanced the cytotoxicity of a combination of 5-fluorouracil (5-FU) and interferon-alpha (IFN) against human colon cancer cell lines in vitro. To test the efficacy of this combination in the clinical setting, patients with locally advanced or advanced gastric carcinoma were treated with the combination of PALA, 5-FU and IFN (PFI). Patients were required to have biopsy-proven disease beyond the scope of surgical resection, measurable disease, no prior chemotherapy, adequate bone marrow, renal and hepatic function, to be fully ambulatory and to have given informed consent. Drug was administered as follows: PALA, 250 mg/m2, 15 min i.v. infusion, days 1, 15, 22, 29, and then weekly; 5-FU, 750 mg/m2 daily x 5 as a continuous i.v. infusion beginning day 2, then at 750 mg/m2 days 16, 23 and 30, then weekly; IFN, 9 MU subcutaneously three times per week beginning day 2. There were 22 patients enrolled. The major toxicities were fatigue and associated neurotoxicity, with acceptable gastrointestinal and haematological toxicities. There was one complete responder (5%) and 3 partial responders (14%); two of these responses were durable (> 3 years). Despite this modest clinical activity, other regimens for advanced gastric cancer such as FAMTX and ELF appear to have greater activity with comparable toxicity.