International prognostic index, type of transplant and response to rituximab are key parameters to tailor treatment in adults with CD20-positive B cell PTLD: clues from the PTLD-1 trial.

Tailoring treatment by patient strata based on the risk of disease progression and treatment toxicity might improve outcomes of patients with posttransplant lymphoproliferative disorder (PTLD). We analysed the cohort of 70 patients treated in the international, multicenter phase II PTLD-1 trial (NCT01458548) to identify such factors. Of the previously published scoring systems in PTLD, the international prognostic index (IPI), the PTLD prognostic index and the Ghobrial score were predictive for overall survival. None of the scoring systems had a considerable effect on the risk for disease progression. Age and ECOG performance status were the baseline variables with the highest prognostic impact in the different scoring systems. Baseline variables not included in the scoring systems that had an impact on overall survival and disease progression were the type of transplant and the response to rituximab at interim staging. Thoracic organ transplant recipients who did not respond to rituximab monotherapy were at particularly high risk for death from disease progression with subsequent CHOP-based chemotherapy. Patients in complete remission after four courses of rituximab and patients in partial remission with low-risk IPI had a low risk of disease progression. We speculate that chemotherapy might not be necessary in this patient cohort.

Prognostic impact of meningeal dissemination in primary CNS lymphoma (PCNSL): experience from the G-PCNSL-SG1 trial.

BACKGROUND: We evaluated the frequency and prognostic impact of meningeal dissemination (MD) in immunocompetent adult patients with primary central nervous system lymphoma treated in a randomized phase III trial. PATIENTS AND METHODS: MD was evaluated at study entry and defined by lymphoma proof in the meningeal compartment detected by at least one of the following methods: cerebrospinal fluid (CSF) cytomorphology, detection of clonal B cells by IgH PCR in CSF or contrast enhancement of the leptomeninges on magnetic resonance imaging (MRI). RESULTS: Data on MD were available in 415 patients, of those, MD was detected in 65 (15.7%): in 44/361 (12.2%) by CSF cytomorphology, in 16/152 (10.5%) by PCR and in 17/415 (4.1%) by MRI. Major patients' characteristics and therapy did not significantly differ between patients with MD (MD+) versus those without MD (MD-). There was a significant correlation of MD with CSF pleocytosis (>5/µl; P < 0.0001), but no correlation with CSF protein elevation (>45 mg/dl). Median progression-free survival was 6.7 months [95% confidence interval (CI) 0-14.5] in MD+ and 8.3 months (5.7-10.8) in MD- patients (P = 0.95); median overall survival was 21.5 months (95% CI 16.8-26.1) and 24.9 months (17.5-32.3), respectively (P = 0.98). CONCLUSION: MD was detected infrequently and had no impact on outcome in this trial.

[Recurrent hypoglycemia and a large intraabdominal tumor in a 61-year-old woman]. / Rezidivierende Hypoglykämien und ein großer intraabdominaler Tumor bei einer 61-jährigen Patientin.

HISTORY AND ADMISSION FINDINGS: A 61-year-old woman was found unconscious by her husband. The emergency doctor detected hypoglycemia (blood glucose 1.7 mmol/l). This was the first such event, the patient had not been known to have diabetes mellitus. At admission the physical examination and the laboratory findings revealed no abnormalities. INVESTIGATIONS: A fasting test was aborted shortly after the start because of the onset of neurological symptoms. An insulinoma was excluded by detecting suppressed levels of insulin and C-peptide. Computed tomography of the abdomen revealed a mesenteric tumour of 9 cm in diameter, which was identified immunhistologically as a grade 1 follicular lymphoma (FL).  After exclusion of endocrinological causes the recurrent hypoglycaemia was diagnozed as part of a paraneoplastic syndrome associated with a non-islet cell tumour hypoglycaemia (NICTH) with a newly diagnosed FL. TREATMENT AND COURSE: Specific medication with the CD20 antibody rituximab (375 mg/m2, once per week for a total of four cycles) was initiated. There were no further episodes of hypoglycaemia. After one year the patient remains free of any symptoms. CONCLUSIONS: After exclusion of any endocrinological reasons for hypoglycemia, differential diagnosis should include NICTH as paraneoplastic syndrome. In rare cases a hematological malignancy may be the underlying disease. The specific treatment of this disease likewise represents the causal treatment of NICTH.

Invasive zygomycosis in patients with graft-versus-host disease after allogeneic stem cell transplantation.

Invasive mold infections are a threat to immunosuppressed patients such as patients with graft-versus-host disease (GVHD) after allogeneic stem cell transplantation (SCT). Up to 10% of SCT recipients develop invasive aspergillosis (IA). Invasive zygomycosis (IZ) may occur during treatment against IA. Here we report 4 SCT patients with GVHD diagnosed with IZ. All patients had received myeloablative hematopoietic SCT and developed chronic GVHD requiring systemic immunosuppression. Underlying diseases were acute lymphocytic leukemia (2), osteomyelofibrosis, and multiple myeloma. All patients had developed pulmonary infiltration that led to initiation of antifungal therapy. Treatment for IA was voriconazole, caspofungin, or itraconazole. Organs involved with zygomycosis were lung, nasal sinus, skin, and kidney. Treatment with liposomal amphotericin and posaconazole was initiated in all patients, and 2 patients also had surgical debridement as well. Despite intensive treatment, no patient survived. IZ is becoming more common in patients with GVHD on successful treatment for IA. Even non-specific symptoms are suspicious in this group of patients and need to be evaluated by vigorous diagnostics. Despite effective antifungals and surgical intervention, the prognosis is grim in patients with active GVHD, as immunoreconstitution is mandatory for successful management.

[Pancytopenia and lymph node swelling. Cardinal symptoms of an unusual differential diagnosis]. / Panzytopenie und generalisierte Lymphknotenschwellung. Leitsymptome einer ungewöhnlichen Differenzialdiagnose.

A 47-year-old woman was admitted to our emergency room because of anemia and acute tonsillitis. She reported recurrent fever and a sore throat. Clinical examination and CT scans showed general lymph node swelling and liver enlargement. In the course of the disease she developed pancytopenia with neutropenic fever, pleuropneumonia, and deep vein thrombosis. The histological examination of a lymph node showed a reactive, EBV-associated lymphadenitis. The examination of the bone marrow showed an activated marrow. The diagnosis of an active EBV infection was established with 2 x 10(6)/ml EBV gene copies in the blood. In addition, systemic lupus erythematosus was diagnosed because of the typical autoantibody constellation and clinical findings. The immunohematological examination showed autoantibodies against the three blood cell compartments. Because of the severe pancytopenia as a result of the EBV- and SLE-associated autoantibodies and despite recurrent infections, we initiated immunosuppressive therapy with low-dose corticosteroids. This therapy resulted in normalization of the blood counts. Anitibody levels and the EBV genome levels became negative.

[Genitourinary manifestations of lymphoma]. / Urogenitale Lymphommanifestationen.

Non-Hodgkin lymphoma (NHL) of the genitourinary system is a very rare disease. Only 1 % of NHL are primary genitourinary lymphomas, most commonly of testicular origin. Secondary infiltration from disseminated lymphoma can be found in all the genitourinary organs. We report on five patients with genitourinary lymphoma who were diagnosed at our department: one case of primary testicular lymphoma and four patients with secondary lymphomatous infiltration of kidney, bladder, prostate and testicles, respectively. The clinical courses are described and the current literature is reviewed. The operation could be restricted to a biopsy for assurance of the diagnosis. However, the mostly radical procedures are performed for curative purposes after an incorrect diagnosis of a malignant organ cancer without even recognizing or considering NHL in the differential diagnosis.

Multivariate analyses of prognostic factors in acute myeloid leukemia: relevance of cytogenetic abnormalities and CD34 expression.

Identification of additional prognostic factors besides karyotype is important for the improvement of the risk adapted treatment strategies in acute myeloid leukemia (AML). The aim of this study was to investigate whether other factors besides karyotype could be used as a prognostic tool in newly diagnosed AML. Biological and disease related established and potential prognostic factors were retrospectively analysed in 124 consecutive AML patients treated between 1993 and 2002 at the University hospital Rostock (Germany). One hundred patients received a potential curative intensive chemotherapy (81%), of whom 28 received an allogeneic HSCT at some point of their treatment course, 17 patients (14%) received palliative therapies and 7 patients (5%) received supportive care only. In patients that received potential curative therapies LDH >or=2000 U/l, WBC >50 GPT/l, CD34 surface expression on the AML blasts, secondary AML, unfavorable karyotype and no allogeneic HSCT at some point of treatment course were associated with unfavorable prognosis. However, in the multivariate risk factor analyses only unfavorable karyotype (p=0.012), CD34 positivity of AML blasts (p=0.046), no allogeneic HSCT (p=0.008) and first diagnosis after 1997 (p=0.025) were independent unfavourable prognostic factors. In conclusion, karyotype and CD34 expression are independent prognostic markers in newly diagnosed AML. Furthermore, receiving an allogeneic HSCT at some point of the treatment course seems to be of benefit for AML patients.

Treatment with campath-1H for relapsed chronic lymphocytic leukemia after allogeneic peripheral blood stem cell transplantation does not abrogate the development of chronic GVHD.

The graft vs. leukemia (GVL) effect is one of the most important factors of anti-tumor activity after allogeneic hematopoetic stem cell transplants (alloSCT). Its effectiveness depends mainly on the tumor biology as well as the tumor burden. Patients with a high tumor burden may not respond to GVL-effect despite otherwise sensitive biology. Campath-1H is known as an effective treatment of chronic lymphocytic leukemia (CLL). Due to its ability to induce profound immunosuppression, it has also been used as part of conditioning regimens before alloSCT. We report a patient, who received campath-1H in combination with docetaxel for treatment of chemotherapy and donor lymphocyte infusion resistant CLL after alloSCT, who developed shortly after discontinuation of treatment with campath-1H severe eosinophilia of the peripheral blood and typical clinical as well as histological signs of cutaneous chronic graft vs. host disease followed by complete clearance of CLL. The clinical course demonstrates the impact of the tumor burden on the GVL-effect, as well as the effectiveness of campath-1H in the presence chemo-resistance in a patient with CLL. Furthermore, the GVL effect was not abrogated by the use of campath-1H.

Treatment of peripheral T-cell lymphomas (PTCL) with high-dose chemotherapy and autologous or allogeneic hematopoietic transplantation.

Peripheral T-cell lymphomas (PTCL) are a rare entity of non-Hodgkin's lymphomas (NHL). Despite the poor outcome after conventional chemotherapy, the impact of high-dose chemotherapy (HDCT) and autologous or allogeneic stem cell transplantation is not well defined in these patients. In a retrospective study, we evaluated the outcome of 15 patients (9 male, 6 female) with PTCL after HDCT with autologous (10 patients) and allogeneic (5 patients) stem cell transplantation between 1996 and 2001 at our department. At the time of transplantation three patients were in second remission, seven patients were in partial remission (PR), and three patients had refractory disease. Two patients were treated with sequential HDCT (cyclophosphamide, adriamycin, vincristine, etoposide, prednisolone, m-CHOEP). The conditioning regimes were heterogeneous. After HDCT ten patients (67%, autologous 7, allogeneic 3) achieved CR, two patients (13%, autologous 2, allogeneic 0) had refractory disease, and three patients (20%, autologous 1, allogeneic 2) died because of toxic side effects before evaluation of response was performed. The median overall survival (OS) was 12 months. The 1-year probability of survival for the autologous and allogeneic groups was 58% and 40%, respectively. At the time of evaluation, six patients are alive and nine patients have died (four severe infection, one late toxicity, two disease progression, and two relapse). Despite the small number of patients in this study, HDCT with autologous or allogeneic hematopoietic transplantation seems to be an effective treatment option that can achieve CR for patients with PTCL. Because of the poor outcome of these patients after conventional chemotherapy, HDCT seems to be a rational option in first-line therapy. Whether it improves survival in these patients should be further investigated.

Progressive interstitial fibrosis of the lung in sclerodermoid chronic graft-versus-host disease.

Sclerodermoid chronic graft-versus-host disease (sGVHD) is a well-known complication in patients with a long history of chronic GVHD. Pulmonary involvement in chronic GVHD presents typically as bronchiolitis obliterans (BO). Pulmonary fibrosis after allogeneic hematopoietic stem cell transplantation (HSCT) is presumed to be caused by the long-term toxicity of the conditioning regimen or the result of lung injury elicited predominantly by viral infections or GVHD. We present two patients with late onset pulmonary fibrosis associated with moderate sGVHD of the skin after HSCT. At the initial diagnosis of chronic GVHD both patients presented with symptoms of interstitial pneumonia. Years later both patients developed moderate to severe interstitial pulmonary fibrosis in association with sGVHD. One patient showed additional clinical and histological signs of BO. While one patient responded to increased immunosuppression including total nodal irradiation (1 Gy), the other patient died due to complications related to pulmonary fibrosis.

High-dose etoposide phosphate and G-CSF mobilizes peripheral blood stem cells in patients that previously failed to mobilize.

Ten consecutive patients in our unit who had failed to mobilize a sufficient stem cell yield after either an initial or several mobilization regimens received high-dose etoposide phosphate (1500-2000 mg/m2) followed by granulocyte colony-stimulating factor (G-CSF; 10 micrograms/kg per day) to stimulate mobilization. Eight of the ten patients were apheresed. A median of 2.1 x 10(6) CD34+/kg (range 0-5.2) was collected. The number of CD34+ cells/microliter peripheral blood (pB) was significantly increased compared to the first-line mobilization [median 13.0 (range 2.68-29) versus median 4.76 (range 1.36-12); P < 0.05]. Besides hematotoxicity and four cases of infection (WHO grade 3), no major side effects were seen. The median duration of neutropenia was short (5 days, range 0-10), which is important in heavily pretreated patients. These results indicate that high-dose etoposide phosphate with G-CSF is safe, well tolerated, and may be effective in peripheral blood stem cell (PBSC) mobilization in patients who had previously failed to mobilize.

Pentoxifylline, cyclosporine A and taurolidine inhibit endotoxin-stimulated tumor necrosis factor-alpha production in rat mesangial cell cultures.

Tumor necrosis factor-alpha (TNF-alpha) is an important mediator in the pathogenesis of glomerular disease. Intrinsic glomerular cells as well as extraglomerular cells have been found as a source of TNF-alpha. Rat glomerular mesangial cells produce TNF-alpha after stimulation with bacterial lipopolysaccharide (0.1, 1.0 and 10 microg/ml) over different times (4, 8, 16 and 24 h). We show that lipopolysaccharide-induced production of TNF-alpha in rat mesangial cell cultures is inhibited by pentoxifylline (50 mg/ml), cyclosporine A (0.1 microg/ml) and taurolidine (100 mg/ml). Inhibition of this production seems to be a promising treatment option for renal disease. Already pentoxifylline and cyclosporine A have been shown to improve different glomerular pathologies. Their in vitro effect on TNF-alpha production shown here might influence this.

The role of the C-terminus of the insulin B-chain in modulating structural and functional properties of the hormone.

Within the scope of structure-function studies on the proteohormone insulin, the role of the C-terminal segment B26-B30 for self-association and receptor interaction was analyzed. Insulin derivatives with modifications in the region B26-B30 were synthesized by trypsin-catalyzed coupling reactions of des-(B23-B30)-insulin with synthetic peptides. The peptides were obtained by Fmoc solid-phase peptide synthesis. Insulins with multiple amino acid-->glycine substitutions were examined to distinguish between the influence of the side chains and the influence of the main chain in positions B27-B30 on the self-association of the hormone. The analogues [GlyB27,B28,B29,B30]insulin and [GlyB27,B28,B30]insulin exhibit relative receptor affinities of 80% and self-associate. The successive extension of [AlaB26]des-(B27-B30)-insulin-B26-amide (relative receptor binding 273%) with amino acids corresponding to the native sequence B27-B30 showed the influence of the length of the B-chain on receptor affinity: the extension by B27-threonine amide reduces receptor binding to 71%, all further prolongations have only small effects on the binding. The effect of the B28-side chain on main-chain conformation, self-association and receptor binding was examined with [XB28]des-(B29-B30)-insulin-B28-amides (X = Phe, Gly, D-Pro). While the glycine and D-proline analogues (relative binding 104 and 143%, respectively) retain the self-association properties typical of insulin, [PheB28]des-(B29-B30)-insulin-B28-amide (relative binding 50%) shows diminished self-association. The backbone-modified insulin derivative [SarB26]des-(B27-B30)-insulin-B26-amide (sarcosine = N-methylglycine) exhibits an unexpectedly high receptor affinity of 1100% which demonstrates that the B26-amide hydrogen of the native hormone is not important for receptor binding.

Proteolyses of a fluorogenic insulin derivative and native insulin in reversed micelles monitored by fluorescence emission, reversed-phase high-performance liquid chromatography, and capillary zone electrophoresis.

The preparation and substrate properties of the fluorogenic insulin derivative N alpha A1-aminobenzoyl-N epilson B29-Tyr(NO2)- insulin are described. This semisynthetic protein intramolecularly quenched by long-range resonance energy transfer between the donor/acceptor pair 2-aminobenzoic acid and 3-nitrotyrosine was used to prove the activity of serine proteases toward substrates of high molecular weight after incorporation in reversed micelles. The proteases investigated, trypsin and alpha-chymotrypsin, were shown to be hydrolytically active in reversed micellar solvent systems stabilized by cetyltrimethylammonium bromide or sodium-1,2-bis(2-ethylhexylcarbonyl)-1- ethane sulfonate. Apart from fluorometric enzyme assays, methods for monitoring proteolyses in reversed micelles were elaborated using either reversed-phase high-performance liquid chromatography or capillary zone electrophoresis. Enzymatic digestions of native insulin by the specific protease trypsin and the less specific protease alpha-chymotrypsin were performed. In contrast to aqueous solution, high but still variable specificity of alpha-chymotrypsin which was dependent on the micellar environment was observed. The results promise further insight into the influence of interfacial environments on enzyme action and a novel approach to enzyme-mediated protein modifications by the use of microstructured solvent systems.

Tumor cell-derived 12(S)-hydroxyeicosatetraenoic acid induces microvascular endothelial cell retraction.

Our previous work demonstrated that the 12-lipoxygenase metabolite of arachidonic acid, 12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE] induced a nondestructive and reversible retraction of cultured endothelial cells. In the current study we tested the hypothesis that tumor cells produce 12(S)-HETE during their interactions with endothelial cells which in turn induces endothelial cell retraction. Coincubation of Lewis lung carcinoma cells or elutriated B16 amelanotic melanoma (B16a) cells but not 3T3 fibroblasts with microvascular endothelial cells (CD3) resulted in a time- and concentration-dependent retraction of the CD3 monolayers as revealed by quantitative binding assays and phase contrast microscopy. Lewis lung carcinoma cell-induced endothelial cell retraction was blocked by specific lipoxygenase inhibitors but not by cyclooxygenase inhibitors, suggesting the involvement of a lipoxygenase metabolite(s). Radioimmunoassay and high-performance liquid chromatography analysis of tumor cell extracts identified 12(S)-HETE as the major lipoxygenase metabolite of arachidonic acid and tumor cell generation of 12(S)-HETE was specifically blocked by a select 12-lipoxygenase inhibitor N-benzyl-N-hydroxy-5-phenyl-pentamide. The identity and stereochemistry of tumor cell-derived 12-HETE was substantiated by gas chromatography-mass spectrometry analysis and chiral phase high-performance liquid chromatography, respectively. Lewis lung carcinoma cell adhesion to CD3 monolayers was accompanied by an enhanced 12(S)-HETE biosynthesis by tumor cells, which paralleled the tumor cell-induced endothelial cell retraction in a cell number-dependent manner. Pretreatment of tumor cells with N-benzyl-N-hydroxy-5-phenylpentamide inhibited both increased 12(S)-HETE biosynthesis and tumor cell-induced endothelial cell retraction. Highly metastatic variants of elutriated B16a cells which had been shown to produce large quantities of 12(S)-HETE induced significant CD3 cell retraction, while low metastatic subpopulations of B16a cells which synthesized no or little 12(S)-HETE did not induce endothelial cell retraction. These results suggest that 12(S)-HETE synthesis during tumor cell-endothelial cell interactions may represent a key contributory factor in cancer metastasis.

1,N6-ethenoadenosine formation, mutagenicity and murine tumor induction as indicators of the generation of an electrophilic epoxide metabolite of the closely related carcinogens ethyl carbamate (urethane) and vinyl carbamate.

Previous studies from this laboratory showed that (i) vinyl carbamate (VC) was much more carcinogenic than ethyl carbamate (EC) and that both carbamates induced the same spectrum of tumors in mice and rats, (ii) adducts of [14C]- or [3H]1,N6-ethenoadenosine and [14C]- or [3H]3,N4-ethenocytidine e were formed in the hepatic RNA of infant male B6C3F1 mice administered [1-14C]ethyl or [1,2-3H]ethyl EC and (iii) VC formed much more of the 1,N6-ethenoadenosine (epsilon Ado) adduct in the hepatic RNA and the 7-(2-oxoethyl)-guanine adduct in the hepatic DNA of mice than did EC. By analogy to the similar results of earlier studies by other investigators on the related carcinogen vinyl chloride, the above data suggested that VC epoxide was a reactive electrophilic metabolite of these carbamates. In the present studies, VC, but not EC, was found to be oxidized by 3-chloroperbenzoic acid to a derivative that reacted with adenosine to form epsilon Ado. Far more of this etheno nucleoside was formed from VC than from EC when these carbamates were metabolized by cofactor-fortified mouse liver microsomes in the presence of adenosine. Sodium diethyldithiocarbamate strongly inhibited these microsomal reactions and the formation of epsilon Ado in the hepatic RNA of mice administered either carbamate. Likewise, the i.p. preadministration of deithyldithiocarbamate markedly inhibited the induction of tumors by single i.p. doses of EC or VC in the livers of infant male B6C3F1 mice and in the livers, lungs and Harderian glands of infant female B6C3F1 mice. This inhibitor also considerably reduced lung tumor induction by VC in adult female A/Jax mice. 2-(2,4-Dichloro-6-phenyl) phenoxyethyl amine, a cytochrome P450 inhibitor, reduced the carcinogenicity of low doses of EC but appeared to increase the carcinogenicity of low doses of VC. The mutagenicity of VC for Salmonella typhimurium TA1535 in the presence of a hepatic activating system was greatly reduced by these inhibitors. The data from all these studies are consistent with the proposal that VC epoxide is an ultimate electrophilic and carcinogenic metabolite of EC and VC in the mouse.