Aims: To investigate the association of acute parvovirus B19 infection with new onset of acute lymphoblastic and myeloblastic leukaemia.
Methods: Cerebrospinal fluid (CSF) samples from patients with acute myelogenous leukaemia (AML) at diagnosis (n = 2) and acute lymphoblastic leukaemia (ALL) at diagnosis (n = 14) were analysed for parvovirus B19 DNA by means of nested polymerase chain reaction. In addition, samples from patients with benign intracranial hypertension (BIH) (n = 10) and hydrocephalus (n = 13) were tested as controls.
Results: Four leukaemia cases were positive—common ALL (n = 2), null cell ALL (n =1), and M7 AML (n = 1)—whereas all controls were negative (Yates corrected χ2 value, 3.97; p = 0.046; odds ratio, 16.92; confidence interval, 1.03 to 77.18). All four patients were significantly anaemic, but none was encephalitic or had evidence of central nervous system leukaemia. In three of these patients, serum tumour necrosis α, interferon γ, interleukin 6, granulocyte–macrophage colony stimulating factor (range, 34.93–3800.06pg/ml), and macrophage chemoattractant protein 1 were detectable. All of these four patients carried at least one of the HLA-DRB1 alleles, which have been associated with symptomatic parvovirus B19 infection.
Conclusion: Erythroid suppression and immune cell proliferation are both associated with B19 infection and may also be important in the pathogenesis of acute leukaemia.
parvovirus B19; acute leukaemia; cytokine; HLA-DR
We previously reported that children in the UKALL XI ALL trial with HLA-DP 1 and -DP 3 supertypes had significantly worse event-free survival (EFS) than children with other DP supertypes. As DP 1 and DP 3 share two of four key antigen-binding amino-acid polymorphisms (aspartic acid84–lysine69), we asked whether Asp84-Lys69 or Asp84 alone were independent prognostic indicators in childhood acute lymphoblastic leukemia (ALL). We analysed EFS in 798 UKALL XI patients, stratified by Asp84-Lys69 vs non-Asp84-Lys69, for a median follow-up of 12.5 years. Asp84-Lys69 was associated with a significantly worse EFS than non-Asp84-Lys69 (5-year EFS: Asp84-Lys69: 58.8% (95% CI (confidence of interval): 52.7–64.9%); non-Asp84-Lys69: 67.3% (63.4–71.2%); 2P=0.007). Post-relapse EFS was 10% less in Asp84-Lys69 than non-Asp84-Lys69 patients. EFS was significantly worse (P=0.03) and post-relapse EFS marginally worse (P=0.06) in patients with Asp84 compared with Gly84. These results suggest that Asp84-Lys69 predicted adverse EFS in the context of UKALL XI because of Asp84, and may have influenced post-relapse EFS. We speculate that this may be due to the recruitment of Asp84-Lys69-restricted regulatory T cells in the context of this regimen, leading to the re-emergence of residual disease. However, functional and molecular studies of the prognostic value of this and other HLA molecular signatures in other childhood ALL trials are needed.
childhood ALL; HLA-DP supertype; DP molecular signature; event-free survival; relapse
Childhood B-cell precursor (BCP) ALL is thought to be caused by a delayed immune response to an unidentified postnatal infection. An association between BCP ALL and HLA class II (DR, DQ, DP) alleles could provide further clues to the identity of the infection, since HLA molecules exhibit allotype-restricted binding of infection-derived antigenic peptides. We clustered >30 HLA-DPB1 alleles into six predicted peptide-binding supertypes (DP1, 2, 3, 4, 6, and 8), based on amino acid di-morphisms at positions 11 (G/L), 69 (E/K), and 84 (G/D) of the DPβ1 domain. We found that the DPβ11-69-84 supertype GEG (DP2), was 70% more frequent in BCP ALL (n=687; P<10−4), and 98% more frequent in cases diagnosed between 3 and 6 years (P<10−4), but not <3 or >6 years, than in controls. Only one of 21 possible DPB1 supergenotypes, GEG/GKG (DP2/DP4) was significantly more frequent in BCP ALL (P=0.00004) than controls. These results suggest that susceptibility to BCP ALL is associated with the DP2 supertype, which is predicted to bind peptides with positively charged, nonpolar aromatic residues at the P4 position, and hydrophobic residues at the P1 and P6 positions. Studies of peptide binding by DP2 alleles could help to identify infection(s) carrying these peptides.
HLA-DPB1; supertypes; BCP ALL; case–control comparison; allele frequency; peptide-binding pockets
Gardner and co-workers advanced the hypothesis that the Seascale leukaemia cluster could have been caused by new mutations in germ cells, induced by paternal preconceptional irradiation (PPI) exposure at the Sellafield nuclear installation. Since evidence has shown that PPI can increase the de novo germline mutation rate in hypervariable minisatellite loci, we investigated the hypothesis that sporadic childhood leukaemia might be associated with an increased parental germline minisatellite mutation rate. To test this hypothesis, we compared de novo germline mutation rates in the hypervariable minisatellite locus, CEB1, in family trios (both parents and their child) of children with leukaemia (n=135) compared with unaffected control families (n=124). The majority of case and control germline mutations were paternal (94%); the mean paternal germline mutation rates of children with leukaemia (0.083) and control children (0.156) were not significantly different (odds ratio, 95% confidence interval: 0.50, 0.23–1.08; P=0.11). There were no significant differences in case and control parental allele sizes, case and control germline mutation progenitor allele sizes (2.74 vs 2.54 kb; P=0.56), case and control mutant allele sizes (2.71 vs 2.67 kb; P=0.90), mutant allele size changes (0.13 vs 0.26 kb; P=0.10), or mutational spectra. Within the limitation of the number of families available for study, we conclude that childhood leukaemia is unlikely to be associated with increased germline minisatellite instability.
childhood leukaemia; minisatellite; germline; mutation
Aims: The epidemiological and pathological features of Hodgkin lymphoma (HL) are complex. The Epstein-Barr virus (EBV) is consistently associated with a proportion of cases, and these cases are thought to represent a distinct aetiological subgroup of HL. The aim of the present analysis was to determine the age and sex specific incidence of EBV associated and non-associated HL, analysed separately, using data derived from a population based study–the Scotland and Newcastle epidemiological study of Hodgkin’s disease (SNEHD). This study also provided a unique opportunity to evaluate accuracy in the current diagnosis and classification of HL.
Methods: SNEHD analysed consecutive cases of HL diagnosed in the study area between 1993 and 1997. Diagnostic biopsy material was retrieved, EBV status of tumours was determined, and histological review was performed.
Results: In total, 622 cases were eligible for the study, and EBV studies and histopathological review were performed on biopsy material from 537 and 549 cases, respectively. Accuracy in the overall diagnosis of HL and classification of nodular sclerosis HL was good, but diagnosis of HL in the elderly and classification of other subtypes was less reliable. One third of classic HL cases were EBV associated, and age specific incidence curves for EBV associated and non-associated cases were distinct.
Conclusions: Comparison of age specific incidence curves for EBV associated and non-associated HL supports the hypothesis that these are two distinct aetiological entities. Accuracy in the diagnosis of HL is generally good, but certain subgroups of cases continue to present diagnostic difficulties.
Hodgkin lymphoma; human herpesvirus 4; epidemiology; histopathological review
treatment-related leukaemia; MLL cleavage and rearrangement; HLH
To investigate whether infections or other environmental exposures may be involved in the aetiology of childhood central nervous system tumours, we have analysed for space–time clustering and seasonality using population-based data from the North West of England for the period 1954 to 1998. Knox tests for space–time interactions between cases were applied with fixed thresholds of close in space, <5 km, and close in time, <1 year apart. Addresses at birth and diagnosis were used. Tests were repeated replacing geographical distance with distance to the Nth nearest neighbour. N was chosen such that the mean distance was 5 km. Data were also examined by a second order procedure based on K-functions. Tests for heterogeneity and Edwards' test for sinusoidal variation were applied to examine changes of incidence with month of birth or diagnosis. There was strong evidence of space–time clustering, particularly involving cases of astrocytoma and ependymoma. Analyses of seasonal variation showed excesses of cases born in the late Autumn or Winter. Results are consistent with a role for infections in a proportion of cases from these diagnostic groups. Further studies are needed to identify putative infectious agents.
British Journal of Cancer (2002) 86, 1070–1077. DOI: 10.1038/sj/bjc/6600228 www.bjcancer.com
© 2002 Cancer Research UK
brain tumours; children; aetiology; infection; seasonal variation; space–time clustering
Durophagous crabs successfully hunt hard-shelled prey by subjecting them to extremely strong biting forces using their claws. Here I show that, for a given body mass, six species of Cancer crabs (Cancer antennarius, Cancer branneri, Cancer gracilis, Cancer magister, Cancer oregonensis and Cancer productus) were able to exert mean maximum biting forces greater than the forces exerted in any other activity by most other animals. These strong biting forces were in part a result of the high stresses (740-1350 kN m(-2)) generated by the claw closer muscle. Furthermore, the maximum muscle stress increased with increasing mean resting sarcomere length (10-18 microm) for the closer muscle of the claws of these six Cancer species. A more extensive analysis incorporating published data on muscle stresses in other animal groups revealed that stress scales isometrically with the resting sarcomere length among species, as predicted by the sliding filament model of muscle contraction. Therefore, muscle or filament traits other than a very long mean sarcomere length need not be invoked in explaining the high stresses generated by crustacean claws.
Mycobacterium bovis has the broadest host range of species in the Mycobacterium tuberculosis complex and is responsible for disease in humans and diverse animal species. We report on genotypic differences at multiple loci among 13 isolates derived from a range of human and animal infections. All isolates were classified as M. bovis by phenotypic analysis but could be subdivided into five distinct genotypes based on polymorphisms at the pncA and oxyR loci, the status of the RD5 deletion region, and the spoligotype pattern. These findings suggest the existence of a spectrum of strains with genotypic characteristics between those of M. tuberculosis and M. bovis.
Previous studies of space–time clustering in childhood leukaemia have produced equivocal and inconsistent results. To address this issue we have used Manchester Children's Tumour Registry leukaemia data in space–time clustering analyses. Knox tests for space–time interactions between cases were applied with fixed thresholds of close in space, < 5 km and close in time < 1 year apart. Addresses at birth as well as diagnosis were utilized. Tests were repeated replacing geographical distance with distance to the Nth nearest neighbour. N was chosen such that the mean distance was 5 km. Data were also examined by a second order procedure based on K-functions. All methods showed highly significant evidence of space–time clustering based on place of birth and time of diagnosis, particularly for all leukaemias aged 0–14 and 0–4 years, and acute lymphoblastic leukaemia (ALL) 0–4 years. Some results based on location at diagnosis were significant but mainly gave larger P -values. The results are consistent with an infectious hypothesis. Furthermore, we found an excess of male cases over females involved in space–time pairs. We suggest this may be related to genetic differences in susceptibility to infection between males and females. These findings provide the basis for future studies to identify possible infectious agents. © 2000 Cancer Research Campaign
clustering; childhood leukaemia; infections
A UK population-based case–control study of Hodgkin's disease (HD) in young adults (16–24 years) included 118 cases and 237 controls matched on year of birth, gender and county of residence. The majority (103) of the cases were classified by Epstein–Barr virus (EBV) status (EBV present in Reed–Stenberg cells), with 19 being EBV-positive. Analyses using conditional logistic regression are presented of subject reports of prior infectious disease (infectious mononucleosis (IM), chicken pox, measles, mumps, pertussis and rubella). In these analyses HD cases are compared with matched controls, EBV-positive cases and EBV-negative cases are compared separately with their controls and formal tests of differences of association by EBV status are applied. A prior history of IM was positively associated with HD (odds ratio (OR) = 2.43, 95% confidence interval (CI) = 1.10–5.33) and with EBV-positive HD (OR = 9.16, 95% CI = 1.07–78.31) and the difference between EBV-positive and EBV-negative HD was statistically significant (P = 0.013). The remaining infectious illnesses (combined) were negatively associated with HD, EBV-positive HD and EBV-negative HD (in the total series, for ≥2 episodes compared with ≤1, OR = 0.45, 95% CI = 0.25–0.83). These results support previous evidence that early exposure to infection protects against HD and that IM increases subsequent risk; the comparisons of EBV-positive and EBV-negative HD are new and generate hypotheses for further study. © 2000 Cancer ResearchCampaign
Hodgkin's disease; Epstein–Barr virus; aetiology; late host exposure model; infectious agents
Hodgkin's disease; HLA-DPB1; HVR; susceptibility; resistance; polymorphic amino acid
AIMS--Tissue fibrosis is a common and serious consequence of chronic inflammation. The mechanism linking these two processes is poorly understood. The present study has utilised a human in vivo model of a delayed type hypersensitivity (DTH) reaction, the tuberculin Heaf reaction, induced by intradermal tuberculin in BCG immunised subjects, to dissect the relation between these two processes. METHODS--Punch skin biopsy specimens were obtained on day 5, day 13 and six to 16 weeks following the tuberculin Heaf test in 18 subjects with grade 3 or 4 responses. Skin biopsy specimens from six subjects served as controls. The specimens were examined using immunohistochemical staining for type 1 procollagen and transforming growth factor-beta (TGF-beta), as well as in situ hybridisation for type 1 procollagen messenger RNA (mRNA). RESULTS--Immunohistochemical analysis revealed increased deposition of TGF-beta in tissue matrix in the biopsy specimens obtained on day 5 following the tuberculin Heaf test. There was also extensive type 1 procollagen staining in the biopsy specimens obtained as early as day 5. Procollagen-1 staining was maximal on day 13, and was present in biopsy specimens from tuberculin Heaf test sites up to eight weeks after the tuberculin inoculation. The type 1 procollagen was localised within cells surrounding areas of inflammatory infiltrate and in perivascular tissues. The presence of new collagen formation was confirmed by in situ hybridisation using oligonucleotide probes for type 1 procollagen mRNA in cells in sections from biopsy specimens obtained on day 13. CONCLUSIONS--These data from a human in vivo model of a DTH response indicate that the immune response is intimately associated with an increase in the production of growth factors and the initiation of a fibrotic response.
The expression of allogenic lymphocyte-activating determinants (LAD) on 25 acute leukaemias has been compared with the expression of cell-surface antigens identified by HLA-DR allo- and xeno-antisera. The close correlation between LAD and DR known to occur on normal lymphocytes was not found in leukaemias. Twenty-two LAD+ leukaemias included 2 DR- cases, whilst 2 LAD- leukaemias were DR+. With the exception of 3 leukaemias all were strongly beta 2 microglobulin+. No correlation was found between the % DR+ cells and the level of lymphocyte stimulation. Separation of leukaemia cells on Ficoll gradients into fractions containing different proportions of DR+ cells did not correlate with LAD expression. Furthermore, antisera to DR antigens only partially blocked leukaemic LAD. The results support the notion that LAD on acute leukaemias are not necessarily associated with or identical to HLA-DR antigens, and that the lymphocyte activating capacity of HLA-DR may be modulated.
Acute leukaemias stimulated proliferative and cell-mediated cytotoxic (CMC) responses in vitro in normal (unprimed) lymphocytes. Proliferation was detected by increases in viable cell counts and [3H]dT incorporation in mixed lymphocyte-leukaemia-cell cultures. CMC detected on cultured cell-line targets (CCL) including K562 was generally much stronger than on fresh leukaemia cells, and correlated with stimulation of [3H]dT uptake in the responding lymphocytes. Leukaemias which were resistant as targets to CMC were able competitively to inhibit CMC on K562, though not as efficiently as blocking by K562 itself. With one leukaemia, blocking of CMC increased as the level of CMC on K562 was amplified by greater numbers of stimulating cells in the sensitization phase. This suggests that in certain cases blocking of effector cells by acute-leukaemia cells may depend upon the state of activation of the effector cells. Lymphocytes from a leukaemia patient in remission, treated with allogeneic leukaemia-cell immunotherapy and stimulated in vitro with immunizing leukaemia cells, developed strong anti-leukaemic CMC. A non-immunized patient's lymphocytes did not respond in this way, despite comparable levels of CMC on K562 in both patients. Dual stimulation of unprimed normal lymphocytes and remission lymphocytes with allogeneic or autologous leukaemias and various cell lines, amplified anti-leukaemic CMC, but did not markedly alter CMC or CCL. These data do not formally exclude the mediation of in vitro-stimulated anti-leukaemic CMC by NK-like cells, but suggest that such effector cells differ qualitatively from NK-like cells detected in the absence of anti-leukaemic CMC.
Plasma prednisolone levels have been measured by radioimmunoassay after oral and rectal administration to healthy volunteers and to patients with idiopathic proctocolitis. The amount of prednisolone absorbed from a 20 mg retention enema given to patients with proctocolitis was about 44% of that absorbed from the same dose orally administered. Adrenocortical response to synthetic ACTH in patients receiving prolonged rectal therapy was either normal, or only slightly impaired, and this may be related to the pattern of steroid absorption rather than to the total amount absorbed.
One hundred and ninety-one adults with acute myelogenous leukaemia were treated with combination chemotherapy consisting of daunorubicin and cytosine arabinoside (Barts III). Sixty-three patients achieved remission and were admitted to one of 3 trials of active immunotherapy: immunotherapy alone, immunotherapy and maintenance chemotherapy or neither of these. All patients had weekly clinical and blood examination and monthly marrow examination. Reinduction chemotherapy was given as soon as relapse was diagnosed in the marrow. The most striking observation was that immunotherapy was associated with easy and repeated reinduction of remission and marked prolongation of survival after first relapse when compared with immunotherapy plus chemotherapy. The possible reasons for this and the value of immunotherapy are discussed in relation to the third trial still in progress which includes 2 maintenance arms, immunotherapy alone and surveillance only.
Immunoglobulin (Fc) receptors were detected on leucocytes from patients with acute myeloid leukaemia (AML) by rosette formation with human cDE/cDE erthyrocytes (HE) sensitized with Rhesus (Rh) antisera (HEA). Of 7 Rh antisera tested, erythrocytes sensitized with anti-d (Gm10) detected the highest numbers of rosette-forming cells (HEA-RFC) in normal and AML leucocyte preparations. Using this assay, HEA-RFC was studied in 22 untreated AML patients and ce assay detected 11-6% lymphocyte and 2-1% granulocyte HEA-RFC in normal peripheral blood. Leucocytes from 16 to 22 AML patients had a similar or lower percentage than normal lymphocyte HEA-RFC, which could be explained by the dilution of peripheral blood leucocytes by poorly or non-rosetting leukaemic blasts. Ten of these 16 patients were diagnosed as having acute myeloblasts leukaemia. Six of the 22 AML patients had high HEA-RFC values of which 5 were diagnosed as having myelomonocytic leukaemia. Cytocentrifuge preparations of HEA-RFC showed that the proportion able to form rosettes was lower in myeloblasts than in monoblasts. Enzyme treatment (pronase), inhibition or simultaneous labelling of surface Ig and Fc receptors showed that the characteristic surface Ig found to AML cells is, at least in part, bound to Fc receptors. The HEA-RFC test described in this paper could be useful in the immuno-diagnosis of myelomonocytic leukaemia.
Thymidine incorporation in vitro by remission lymphocytes from a total of 6 patients with acute myeloid leukaemia (AML) was measured following stimulation by autochthonous and allogeneic AML blasts and cell lines. The early peak response to autochthonous blasts in 2 of these patients (48-72 h) is consistent with the concept of a population of lymphocytes pre-immunized to antigens carried by the blasts. Although stimulation in one patient was increased in the presence of more stimulating (S) blasts than responding (R) lymphocytes, positive responses in other tests were obtained at an S : R ratio of 1 : 1-5. When different methods of treatment of the stimulating autochthonous blasts were compared with untreated cells, mitomycin C gave the highest stimulation indices 2 out of 3 tests. Tissue culture medium in which autochthonous blasts had been incubated for 3-5 days failed to stimulate either remission lymphocytes alone, or combined cultures of lymphocytes with autochthonous or allogeneic blasts, suggesting that mitogenic factors released from autochthonous blasts are not responsible for lymphocyte stimulation. Treatment of autochthonous or allogeneic AML blasts with glycine-HC1(pH 3-0) to remove putative "blocking" factors failed to increase the stimulatory capacity of the leukaemic blasts.
Leucocytes from normal individuals and from patients with acute myeloid leukaemia (AML) in remission receiving active immunotherapy with allogeneic AML blasts (AML-I) were cultured for 6 days with AML-I blasts, Burkitt's lymphoma cells (BL) or lymphoblastoid cells (LCL). The leucocytes were then tested for cell-mediated cytotoxicity (CMC) against 51Cr-labelled AML-I, BL or LCL target cells. There was no substantial difference in the CMC of leucocytes from patients and normals cultured without stimulation, and tested against AML-I, BL or LCL targets. Patient's leucocytes stimulated in vitro with AML-I had a greater frequency of positive CMC responses against AML-I, BL and LCL than normal individuals. The results suggest that co-cultivation of leucocytes with AML-I blasts reactivates memory cytotoxic leucocytes in AML patients receiving immunotherapy and that this test may be useful in measuring the effectiveness of immunotherapy.
Comparison of DQA1 and DQB1 alleles in 60 children with common acute lymphoblastic leukaemia (c-ALL) and 78 newborn infant control subjects revealed that male but not female patients had a higher frequency of DQA1*0101/*0104 and DQB1*0501 than appropriate control subjects. The results suggest a male-associated susceptibility haplotype in c-ALL and supports an infectious aetiology.
Infection has long been suspected as a possible factor in the aetiology of leukaemia and lymphoma. If seasonal variation in the onset of disease could be shown in any of the diagnostic subgroups of leukaemia or lymphoma, this would provide supportive evidence of an aetiology linked to exposure to infection. All cases in the Manchester Children's Tumour Registry (aged 0-14 years at diagnosis) with acute lymphoblastic leukaemia (ALL), acute non-lymphocytic leukaemia (ANLL), Hodgkin's disease (HD) or non-Hodgkin lymphoma (NHL) between 1 January 1954 and 31 December 1996 were included in an analysis of seasonal variation in the month of first symptom and the month of diagnosis. Cases of common acute lymphoblastic leukaemia (c-ALL) diagnosed from 1979 onwards were also analysed separately. The groups considered for analysis were: all cases of ALL (n = 1070), ALL diagnosed between 18 and 95 months of age (n = 730), ALL diagnosed over 95 months of age (n = 266), c-ALL (n = 309), ANLL (n = 244), all infant acute leukaemias (ALL and ANLL under 18 months; n = 107), HD (n = 166) and NHL (n = 189). Using the Edwards method, both c-ALL and HD demonstrated significant seasonal variation (P = 0.037 and 0.001 respectively) in date of first symptom, with peaks occurring in November and December respectively. Using this method, no indication of seasonal variation was found in the other diagnostic groups for date of first symptom or in any of the diagnostic groups for date of diagnosis. For comparison with a previous study, a further analysis based on date of diagnosis for all ALL cases, using summer-winter ratios, showed a significant summer excess. These results provide supportive evidence for an infectious aetiology for c-ALL and HD, and possibly for all ALL, which warrants further investigation.