Biomedical subjects
Publications and source records attributed to F Zintl.
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We report a patient who developed Epstein-Barr virus (EBV)-induced transverse myelitis 19 months after unrelated bone marrow transplantation (BMT). The disease was diagnosed by physical examination, serologic determinations, EBV-specific polymerase chain reaction in peripheral blood lymphocytes and cerebrospinal fluid, and characteristic magnetic resonance imaging scan of the spine. The patient was treated with ganciclovir and cytomegalovirus (CMV) hyperimmune globulin. He gradually improved and recovered completely within 4 weeks. This case suggests that ganciclovir and CMV hyperimmune globulin appear to be effective for the treatment of EBV-induced transverse myelitis in immunocompromised patients following BMT.
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Cyclosporin A (CsA) can induce graft-versus-host disease (GVHD) following autologous bone marrow transplantation (ABMT) and autologous peripheral blood stem cell transplantation (APBSCT) in adults. We investigated whether GVHD can be induced following ABMT and APBSCT in childhood, and which cells are involved in the pathogenesis of this syndrome. We conducted a prospective study of 20 children and adolescents with hematological malignancies receiving CsA after ABMT and APBSCT. Skin biopsies were obtained on day 21 after transplantation or in the event of a rash. Immunophenotypic analysis of peripheral blood lymphocytes was performed on days 14, 21, 28 and 60 after transplantation. Clinical GVHD of the skin, confirmed by histological criteria, occurred in five patients. Five patients had no clinical GVHD but had acute GVHD alterations on routine skin biopsy. In all 10 patients with a positive skin biopsy for GVHD, CD4+ lymphocytes were the predominant cells in the epidermis. Immunophenotypic analysis of peripheral blood lymphocytes revealed a significantly increased CD4/CD8 ratio in patients with a positive skin biopsy (P < 0.01). Our findings indicate that it is possible to induce acute GVHD following ABMT and APBSCT in childhood. In addition, CD4+ lymphocytes play an important role in the pathogenesis of CsA-induced GVHD.
In this report we review current studies concerning the RB-1 gene expression in acute leukemias. The RB-1 gene was analyzed in several studies by protein-, RNA and DNA-techniques in acute lymphoblastic leukemia (ALL) as well as in acute myelogenous leukemia (AML). The frequency of RB-1 inactivation in ALL-patients ranged between 30% and 64% in several studies. Structural abnormalities of the RB-1 gene were reported in 18% of ALL-patients and in 27% of Philadelphia chromosome-positive ALL, respectively. The proportion of AML-patients with absent RB-1 protein expression ranged between 19% and 55%. Structural RB-1-abnormalities in AML were predominantly reported in leukemias with monocytic differentiation. Furthermore, the prognostic value of an abnormal RB-1 gene expression was also estimated in some studies. In childhood ALL RB-1 inactivation was reported to have prognostic significance while in contrast, in another study on adults no prognostic value of RB-1 was found. In 4 out of 5 documented studies AML-patients with RB-1 inactivation generally had a poorer prognosis. In conclusion, RB-1 inactivation is frequently observed in acute leukemia. The prognostic value of low RB-1 expression is controversial but the majority of published studies found low RB-1 expression to be a negative prognostic predictor, in acute leukemia.
Immunohistochemistry was used to analyze samples of 40 newly diagnosed childhood acute lymphoblastic leukemias (ALL) for their expression of cyclins (D1, E, A), cyclin-dependent kinases (cdk2, cdk4) and tumor-suppressor genes (pRb, p16INK4A), in order to discover whether or not the expression of these various proteins may be of prognostic relevance for the survival of children with ALL. Patients with ALL who were strongly positive for cyclin D1 had a lower probability of remaining in first continuous remission than ALL patients who were negative or weakly positive for this trait. There was also a significant correlation between expression of cyclin D1 and frequency of recurrence. For cyclin E and cyclin A, in contrast, there was no difference in the duration of relapse-free-intervals or the frequency of recurrence in patients. Children with cdk4-positive ALL had a lower probability of remaining in first continuous remission than children with cdk4-negative ALL. No prognostic relevance was found for cdk2. Patients with ALL who expressed pRb had a higher probability and patients who expressed p16 a lower probability of remaining in first continuous remission, but the results were not statistically significant. This investigation demonstrated that cyclin D1 and cdk4 were the most important prognostic factors for children with ALL, and that the combination of them showed the strongest prognostic relevance.
PATIENTS AND RESULTS: Prior to bone marrow transplantation (BMT), at the end of the conditioning phase, we found in 42 investigated children with malignant diseases subnormal lowered plasma- and blood selenium levels. Parallel to the diminished selenium status the plasma glutathione peroxidase activity (Gpx) was not reduced as it is in selenium deficiency, but markedly elevated and probably reflecting cytolytic processes. In the group of combined conditioning (fractionated total body irradiation plus chemotherapy) we found significantly more elevated plasma Gpx values in comparison to the only-chemotherapy group. The renal selenium excretion was elevated during the whole observation and could be caused by disturbed tubular function. CONCLUSION: We conclude, that in the situation of BMT a selenium substitution in a dosage of at least 1 to 2 micrograms Se/kg/d is necessary. Patients' selenium status should be monitored by analyses of plasma- and blood selenium contents.
In this investigation, untreated non-B-type acute lymphoblastic leukemia (ALL) of 104 children was analyzed using immunocytochemistry for expression of protein kinase C, proto-oncogene products (Fos, Jun, Ras) and resistance-related proteins (topoisomerase II, P-glycoprotein, glutathione S-transferase-pi, metallothionein, dihydrofolate-reductase, thymidylate-synthase). The aim of the analysis was to find out whether combining those factors with the most important clinical prognostic factor (blast cell count) can improve the prognostic value (relapse-free interval). Univariate analysis shows that protein kinase D (PKC), Fos, P-glycoprotein (P-170) and glutathione S-transferase-pi (GST-pi) are significant prognostic factors independent of blast cell count (PBC) for the relapse-free intervals of children with ALL. The presence of the proteins Fos, PKC, P-170 and GST-pi was not independent within the patient population. The multivariate analysis showed that in combination with PBC and PKC, both P-170 and GST-pi have only limited prognostic influence. Combining the factors PKC, Fos and GST-pi as a categorical variable showed that this variable is a strong prognostic factor in addition to PBC.
The knowledge of renal function in the course of BMT is poor. We prospectively investigated glomerular and tubular function in 42 children who underwent BMT because of malignancy. Seventeen children were transplanted autologously. Investigations were performed before and immediately after the conditioning regimen. Inulin and creatinine clearance, albuminuria, urine excretion of alpha 1-microglobulin, beta-N-acetylglucosaminidase, alanine-aminopeptidase, intestinal alkaline phosphatase, and Tamm-Horsfall-Protein as well as sodium- and phosphatreabsorption were measured. The patients were classified regarding use of total body irradiation (tTBI) in the conditioning regimen. BEFORE CR: Glomerular filtration rate (GFR) was not influenced by the underlying diagnosis or previous treatment. Mean GFR was elevated compared with the reference group. Microalbuminuria was elevated in 15% of patients, and mean levels were higher than in the reference group. Proximal tubular dysfunction was indicated by an elevated excretion of alpha 1-MG in 54%, of beta-NAG in 66%, of AAP in 40%, and of IAP in 47%. Fractional sodium excretion was abnormal in 21%, phosphate reabsorption in 5% and THP-excretion in 7% of the patients. AFTER CR: Creatinine clearance was not affected by CR. After CR alpha 1-MG, beta-NAG, FENa, AAP, and IAP were increased compared with values before CR. TP/Clcr was decreased. Excretion of THP was not altered by CR. In patients without fTBI there was a greater increase in alpha 1-MG excretion and decrease in phosphate reabsorption after CR compared with patients conditioned with fTBI. We conclude that significant proximal tubular dysfunction is present in about 50-60% of patients before and in nearly all alter CR. Distal tubular function was less severely affected. Severity of nephrotoxicity after CR did not correlate with pre-existing abnormalities.
The effect of cranial irradiation on possible therapy-induced morphological central nervous system (CNS) side effects of children cured from acute lymphoblastic leukemia (ALL) is controversially discussed. In a retrospective multicenter study, 118 former ALL patients in first continuous remission were investigated using cranial computerised tomography (CCT) or magnetic resonance imaging (MRI) scans to evaluate CNS related impairments. Corresponding to the different kinds of CNS prophylaxis, the patient sample was divided: group A (n = 39) receiving intrathecal methotrexate (ITMTX) and systemical medium-high-dose methotrexate (SMHDMTX), group B (n = 41) cranial irradiated (in mean 16.8 Gy) and administering ITMTX and SMHDMTX, group C (n = 38) irradiated (in mean 17.1 Gy) and getting ITMTX. Pathologic scans showed atrophy, leukoencephalopathy, calcifications or grey matter changes. These findings were compared with the neuropsychological test results. Abnormal MRI or CCI scans were found in 61/118 patients (51.7%). Fifteen belonged to group A (38.5%), 23 to B (56.1%) and 23 to C (60.5%). Patients with definite CNS changes show reduced neuropsychological test results. The prevalence of brain alterations seems to appear twice increased after lengthening the posttherapeutic interval in irradiated patients as in nonirradiated patients. Irradiated patients as an age younger than 2 years at diagnosis may show a lower prevalence for developing CNS alterations. CNS alterations are not sex-related. Children treated with cranial irradiation in combination with SMHDMTX and/or ITMTX were at greater risk of developing morphological brain alterations than patients with chemotherapy alone. These alterations are partly correlated with reduced neuropsychological performances and seem to stay with a longer posttherapeutic interval.
Autologous peripheral blood stem cells, obtained by CD34+ stem cell selection, are being used with increasing frequency for transplantation in patients with neuroblastoma. Here, we examined the surface membrane antigens of neuroblastoma cells with a panel of hematopoietic monoclonal antibodies (mAbs), including anti-CD34 mAbs, by flow cytometric analysis. We found stronger binding of anti-CD34 mAbs to clonogenic, less differentiated, non-adherent neuroblastoma cells than to adherent neuroblastoma cells. Moreover, the majority of neuroblastoma cell lines shared hematopoietic-associated antigens with all blood cells. Because of these cross-reactions, especially found with the anti-CD34 mAbs 12.8 and ICH3, we have demonstrated that there is a potential risk of cell harvest contamination by circulating neuroblastoma cells during CD34+ stem cell selection.
Seventy-two samples with initial and 23 samples with relapsed childhood acute lymphoblastic leukaemia (ALL) were investigated for apoptotic index (AI) and for Fas expression. AI was determined by DNA-fragmentation using deoxynucleotidyl terminal transferase and Fas expression by immunocytochemistry. bcl-2 mRNA expression was measured in 50 initial and 20 relapsed ALL. The patients were discriminated in groups with low and high AI, Fas-protein expression and bcl-2mRNA expression by the mean value. AI was higher in relapsed than in initial ALL. The mean survival was significantly higher in patients with low AI (p= 0.034). This was also true for the relapse-free interval, however, this result was borderline significant (p= 0.064). AI was directly correlated with expression of Fas and inversely correlated with bcl-2 mRNA expression. These results suggest that Fas and - with limitations - bcl-2 may influence the apoptotic process in childhood ALL and that enhanced apoptotic activity predicts poor prognosis.
Ex vivo production of interleukin 10 (IL-10) and interferon-gamma (IFN-gamma) was investigated in patients with (n = 5) or without (n = 5) chronic graft-versus-host disease (cGVHD) after allogeneic BMT. Patients were matched for time after transplantation and type of transplant. Anti-CD3-induced IL-10 production in MNCs isolated from patients with cGVHD (range/median: 26-650 pg/10(6) MNC; 400 pg/10(6) MNC) was significantly reduced compared to patients without cGVHD (646-2662 pg/10(6) MNC; 1314 pg/10(6) MNC; P < 0.05) or healthy controls (679-6361 pg/10(6) MNC; 3054 pg/10(6) MNC, P < 0.01). In vitro inhibition of IL-10 by an anti-IL-10 monoclonal antibody enhanced the release of IFN-gamma by anti-CD3-stimulated MNCs from 354 +/- 34 pg/10(6) MNCs to 899 +/- 61 pg/10(6) MNCs. Thus, low IL-10 production may cause high IFN-gamma release. In anti-CD3-activated MNCs obtained from patients with cGVHD IFN-gamma production was significantly increased (324-3331 pg/10(6) MNC; 1849 pg/10(6) MNC) compared to healthy donors (127-900 pg/10(6) MNC; 305 pg/10(6) MNC P < 0.01). In addition, median IFN-gamma release by anti-CD3-activated MNCs obtained from patients without cGVHD (464 pg/10(6) MNC) was about five-fold lower than in patients with cGVHD. In contrast to cytokine production, the differential leukocyte count (percentages of monocytes, T cells and CD4/CD8 ratio) was essentially the same both in patients with or without cGVHD. Thus, a different activation of Th-1 and Th-2 cells by anti-CD3 may be responsible for the deviant cytokine productions in patients with cGVHD. In conclusion, we observed significantly decreased IL-10 production in patinets with cGVHD and an increased median IFN-gamma secretion, which may contribute to the altered cytokine production after allogeneic BMT leading to cGVHD. Thus, supplementing IL-10 may become a new strategy for preventing cGVHD.
IL-10 plays an important role in the control of immune reactions during systemic infection. Here, IL-10 serum levels were investigated in patients after BMT. The IL-10 levels correlated with the clinical course of the patients and with serum levels of C-reactive protein (CRP) and neopterin (NP). A total of 26 patients with AML (7), ALL (12), CML (2), NHL (3) and multifocal Ewing's sarcoma (2) had received autologous (10) or allogeneic (16) BMT from related (9) or unrelated donors (7). Routine serum samples were obtained prior to BMT and at days 46 and 100 after BMT. However, in patients with severe complications additional samples were drawn at individual points in time. Prior to BMT, IL-10 serum levels were not detectable in 24/24 patients. Post-BMT, 11 patients developed elevated IL-10 levels, of these eight died of complications (DOC), whereas only one of 15 patients with undetectable IL-10 died of complications, indicating that high IL-10 levels were significantly correlated with severe life-threatening complications (chi2, P < 0.01). To determine the pathomechanism and role of the increased IL-10 levels, they were correlated to the respective NP and CRP serum concentrations. CRP and NP concentrations were found significantly elevated in patients with detectable IL-10, indicating a severe acute phase reaction associated with macrophage activation. In conclusion, high IL-10 serum levels in patients after BMT were significantly associated with fatal outcome. Since IL-10 is a strong suppressor of T cell immunity, high IL-10 production in patients with severe complications such as septic shock or GVHD > grade II after BMT might lead to functional immunodeficiency contributing to the poor prognosis of these patients.
Herpes zoster (HZ) is one of the most common complications after bone marrow transplantation (BMT) in children. Apart from treatment with antiviral drugs, effective prevention by active immunization with varicella-zoster virus (VZV) appears to be possible. In this study 15 patients were vaccinated with a live attenuated VZV vaccine (Varilrix) 12-23 months after BMT. The vaccine was well tolerated without adverse reactions. Chickenpox or HZ were not observed for up to 2 years after immunization. Eight out of nine seronegative patients seroconverted and in six virus-specific IgG could still be demonstrated 2 years later. The incidence of VZV diseases in 133 non-immunized children after BMT was 26.3%. Infections usually occurred within 18 months after BMT.
The murine anti-neuroblastoma monoclonal antibodies 15/7 and 19/1/4 should be tested for specific radiolocalization of neuroblastoma by immunoscintigraphic imaging of this tumour growing in mice. Radioiodination of both antibodies was done by chloramine-T method resulted in an immunoreactivity of 75%. The calculated specific activity varied from 51.1 to 126.2 kBq/microgram IgG. In each case, about 500 kBq of labeled antibodies were intraperitoneally injected into human neuroblastoma (SK-N-MC, SK-PN-DW and IMR 5) xenografted severe complete immunodeficient (SCID) mice. Whole-body scintigraphy was performed daily by a scintiscanner to localize the tumour site. After last scanning principal organs were removed and their I-131-uptake was determined by measuring the impulse rate. The best scintigrams were done with I-131-19/1/4 at the second day after antibody injection. Radioconjugates were accumulated at highest in the tumour at the third day after application of 15/7 and 19/1/4 with a tumour uptake of 0.4 and 2.2 per cent of injected dose per gram (%ID/g), respectively. The 15/7-moAbs was accumulated approximately 9-fold higher in the SK-N-MC and SK-PN-DW grafts than in principal organs, whereas the tumour/non-tumour-ratio of the 19/1/4 moAb was 3:1. The results indicate the efficacy of these two neuroblastoma antibodies in radiolabelling and their usefulness for tumour imaging of neuroblastoma engrafted SCID-mice.
Initial and relapsed childhood acute lymphoblastic leukemias (ALL) were investigated for mutations and expression of deoxycytidine kinase (dCK). dCK mediating toxicity of 1-beta-D-arabinofuranosylcytosine was analyzed semiquantitatively by RT-PCR and by single-strand conformation polymorphism. We found that patients with initial ALL experienced more frequently a relapse if dCK expression was low or absent. No mutations were found in initial and relapsed ALL.
Thirty-eight children with initial and 25 children with relapsed lymphoblastic leukemia (ALL) were analyzed for the expression of LRP (lung resistance protein or p110 major vault protein) using immunocytochemistry. LRP expression was found in 18 of 38 (47%) children with initial ALL and in 17 of 25 (68%) children with relapsed ALL. Children with initial ALL and without LRP expression had significantly longer relapse-free intervals than patients with LRP expression.