Allogeneic stem-cell transplantation in patients with refractory acute leukemia: a long-term follow-up.

We examined retrospectively 44 patients with refractory acute leukemia (acute myeloid leukemia (AML)/acute lymphoblastic leukemia=25/19) who underwent allogeneic transplantation at our center between 11/1990 and 04/2004. The median leukemic blasts was 25% and age 28 years (range, 3-56). Twenty-one patients had untreated relapse, 13 failed reinduction, eight in partial remission and two aplastic. Conditioning was myeloablative using cyclophosphamide, busulfan, total-body irradiation and etoposide (Bu/Cy/VP, n=22; TBI/Cy/VP, n=17; others, n=5) followed by marrow or peripheral blood transplant (n=23/21) from unrelated or related donors (n=28/16). All patients had graft-versus-host disease (GVHD) prophylaxis with cyclosporin and methotrexate. One patient experienced late graft failure. Severe acute-GVHD and chronic-GVHD appeared in eight and 14 patients, respectively. Thirteen patients (30%) remain alive after a median of 25.3 months (range, 2.4-134.1); with 31 deaths, mostly from relapse (n=15) and infections (n=12). Overall survival (OS) and progression-free survival (PFS) at 5 years was 28 and 26%, respectively. OS and PFS were significantly better with blasts < or =20% and time to transplant < or =1 year while transplant-related mortality was less with the use of TBI. We conclude that patients with refractory leukemia can benefit from allogeneic BMT, especially with < or =20% marrow blast.

Reduced-toxicity conditioning with treosulfan, fludarabine and ATG as preparative regimen for allogeneic stem cell transplantation (alloSCT) in elderly patients with secondary acute myeloid leukemia (sAML) or myelodysplastic syndrome (MDS).

We investigated a dose-reduced conditioning regimen consisting of treosulfan and fludarabine followed by allogeneic stem cell transplantation (SCT) in 26 patients with secondary AML or MDS. Twenty patients were transplanted from matched or mismatched unrelated donors and six from HLA-identical sibling donors. The median age of the patients was 60 years (range, 44-70). None of the patients was eligible for a standard myeloablative preparative regimen. No graft-failure was observed, and leukocyte and platelet engraftment were observed after a median of 16 and 17 days, respectively. Acute graft-versus-host disease (GvHD) grade II-IV was seen in 23% and severe grade III GvHD in 12% of the patients. No patients experienced grade IV acute GvHD. Chronic GvHD was noted in 36% of the patients, which was extensive disease in 18%. The 2-year cumulative incidence of relapse was 21%. The relapse rate was higher in patients beyond CR1 or with intermediate two or high risk MDS (P = 0.02). The treatment-related mortality at day 100 was 28%. The 2-year estimated overall and disease-free survival was 36-34%, respectively. No difference in survival was seen between unrelated and related SCT.

Comparable results in patients with acute lymphoblastic leukemia after related and unrelated stem cell transplantation.

We report the results of 84 patients with ALL after related (n = 46) or unrelated (n = 38) allogeneic SCT. Mean recipient age was 23 years (range: 1-60) and median follow-up was 18 months (range: 1-133). Forty-three patients were transplanted in CR1; 25 in CR2 or CR3; four were primary refractory; four in PR; eight in relapse. The conditioning regimen consisted of TBI/VP16/CY (n = 76), TBI/VP16 (n = 2), TBI/CY (n = 2), Bu/VP16/CY (n = 4). The OS at 3 years was 45% (44% unrelated, 46% related). Univariate analysis showed a significantly better OS for patients <18 years (P=0.03), mismatched sex-combination (P = 0.03), both with a stronger effect on increasing OS after unrelated SCT. Factors decreasing TRM were patient age <18 years (P = 0.004), patient CMV-seronegativity (P = 0.014), female recipient (P = 0.04). There was no significant difference in TRM and the relapse rate was similar in both donor type groups. Multivariate analysis showed that factors for increased OS which remained significant were mismatched sex-combination (RR: 0.70,95% CI: 0.51-0.93, P = 0.015), patient age < 18 years (RR: 0.66, 95% CI: 0.47-0.93, P = 0.016). A decreased TRM was found for female patients (RR: 0.56, 95% CI: 0.33-0.98, P=0.042), negative CMV status of the patient (RR: 0.57, 95% CI: 0.36-0.90, P = 0.015). Unrelated stem cell transplantation for high-risk ALL patients with no HLA-compatible family donor is justifiable.

A postulated phylogenetic tree for the human apolipoprotein B gene: unpredicted haplotypes are associated with elevated apo B levels.

Using published data on seven polymorphic sites in the human apolipoprotein B (apo B) gene, it is possible to postulate a model phylogenetic tree for this gene, covering the time since the divergence of human beings from other primates. This simple model assumes no obligatory recombination events or multiple occurrences of the same mutation. This model was tested in two samples of Swedish individuals consisting of 143 young, myocardial infarction patients and 90 healthy, age-matched, control individuals. All the haplotypes postulated in the simple model were observed unequivocally. However, in addition, three unpredicted haplotypes were unambiguously observed and a further nine, much rarer haplotypes were deduced to occur in these samples. The frequencies of the haplotypes postulated in the model do not differ between the patient and control samples, however most of the unpredicted haplotypes occur more frequently in the patient group than in the controls. Two of these unpredicted haplotypes, defined by the combination of the Antigen group (a) epitope and the presence of the XbaI cutting site, were associated with raised serum apo B levels in the control group and significantly elevated levels in the patient group. We propose that these observations explain in part the consistent association reported between the XbaI polymorphic site in the apo B gene and levels of plasma lipids.

Apolipoprotein B gene polymorphisms are associated with lipid levels in men of South Asian descent.

Three polymorphic sites of the apolipoprotein B gene - the insertion/deletion signal peptide, XbaI and EcoRI sites - were examined in a sample of 107 healthy men and in 46 men with evidence of coronary heart disease selected from a large population survey of South Asians aged 40-69 in London, U.K. There were no significant differences in allele frequencies between cases and controls. Frequencies of the ins (insertion) and X- (absence of XbaI cutting site) alleles were higher in South Asians than in Europeans studied previously (South Asians versus Europeans ins: 0.80 vs. 0.68, P less than 0.025; X-: 0.71 vs. 0.47-0.56, P less than 0.001). The del allele was associated with higher levels of total cholesterol (P less than 0.05) and the X+ allele with lower levels of HDL cholesterol (P less than 0.05), and thus both polymorphisms were associated with differences in the ratio of HDL cholesterol to total cholesterol (ins/del, P less than 0.01; XbaI, P less than 0.001). Mean waist-hip girth ratio was lower in the 10 men homozygous for the X+ allele than in the 42 men with X-/X+ and 55 men with X-/X- genotypes; the means (+/- SEM) were 0.92 +/- 0.02, 0.97 +/- 0.01 and 0.96 +/- 0.01 respectively (P = 0.03). These data suggest that genetic variation in linkage disequilibrium with the XbaI and ins/del polymorphisms of the apo B gene contributes to the determination of total cholesterol and HDL cholesterol levels and possibly to obesity in South Asians.

Treatment of mycotic infections after haemopoietic progenitor cell transplantation with liposomal amphotericin-B.

115 patients undergoing allogeneic or autologous bone marrow or peripheral blood stem cell transplantation were treated empirically or for documented fungal infection with liposomal amphotericin-B in doses up to 10mg/kg bodyweight for a duration up to 61 days. The therapy was excellent tolerated and clinical side effects occurred in only eight patients. The drug had to be withdrawn in one episode. A significant influence of liposomal amphotericin-B on laboratory parameters was not observed. Creatinine increased under therapy from a median base point of 1,0 (0,2-3,5) mg/dl to the upper normal value of 1,4 (0,4-4,2) mg/dl. Heavy increases of creatinine as well as of bilirubin, OT and PT were mostly associated with GvHD or regimen related toxicity. Considering the high-risk state of the patients the overall response rate was favourable with 62,9%. However, despite administration of liposomal amphotericin-B culture-proven mycoses were associated with a high morbidity (93,3%). Only one of fourteen patients was cured from Candida lambica septicaemia. We conclude that the antimycotic therapy with liposomal amphotericin-B has a low incidence of side effects. This should, considering the high mortality of fungal infections in BMT recipients, encourage investigators to perform dose escalating studies against the conventional formulation.

Stem cell mobilisation with 16 microg/kg vs 10 microg/kg of G-CSF for allogeneic transplantation in healthy donors.

We compared two doses of recombinant human granulocyte-stimulating factor (G-CSF) for stem cell mobilisation in 90 healthy donors for allogeneic stem cell transplantation in a retrospective analysis. Group I (n = 46) received 10 microg/kg G-CSF (filgrastim) given as 5 microg/kg twice daily, and group II (n = 44) received 16 microg/kg, given as 8 microg/kg twice daily with a 12-h interval. The groups were well-balanced for age and body-weight. G-CSF application was performed on an out-patient basis, and leukapheresis was started in all donors on day 5. The most frequent side-effects of G-CSF were grade I/II, bone pain, headache and fatigue in both groups, whereas grade III of bone pain, headache and fatigue occurred in the 2 x 8 microg/kg group only. One serious non-fatal event with non-traumatic spleen rupture occurred in the 2 x 5 microg/kg group. The CD34(+)cell count in the first apheresis of all donors was 5.1 x 10(6)/kg donor weight (range, 1.5-19.3). The CD34(+) cell harvest was higher in the 2 x 8 microg/kg group than in the 2 x 5 microg/kg group (7.1 x 10(6)/kg vs 4.9 x 10(6)/kg; P = 0.09). The target of collecting >5.0 x 10(6) CD34(+) cells/kg donor weight with one apheresis procedure was achieved in 45% of group I and in 61% of group II, respectively. Administering G-CSF at a dosage of 8 microg/kg twice daily leads to a higher CD34(+) cell yield than a dosage of 2 x 5 microg/kg, but is associated with increased toxicity and higher cost.

Comparison of total body irradiation vs busulfan in combination with cyclophosphamide as conditioning for unrelated stem cell transplantation in CML patients.

We compared fractionated total body irradiation (12 Gy)/cyclophosphamide (120 mg/kg) with busulfan (16 mg/kg)/cyclophosphamide (120 mg/kg) as preparative therapy in unrelated donor stem cell transplantation of CML patients. Fifty patients with CML (1.CP = 46; aP = 4) and a median age of 36 years (range 16-52) were enrolled in this sequential trial between 1994 and 1999. In both groups patients were well balanced with respect to age, disease status, stem cell source and CMV status. All patients received standard doses of cyclosporin A, methotrexate and anti-thymocyte globulin (ATG) as GVHD prophylaxis. No graft failures occurred in either group. The median day of leukocyte engraftment was earlier in the Bu/Cy than in the TBI/Cy group (day 15 vs 17; P = 0.006). The incidence of grade II-IV GVHD was 40% in the TBI/Cy and 36% in the Bu/Cy group, whereas severe grade III/IV GVHD was only observed in 12% of patients in both groups. The incidence of chronic GVHD (limited and extensive) at 1 year was higher in the Bu/Cy arm (65% vs 30%; P = 0.02). More toxicity grade I/II of the liver (88% vs 44%; P = 0.002) and more hemorrhagic cystitis (32% vs 8%; P = 0.02) were observed in the Bu/Cy regimen. Seven relapses in the TBI and no relapse in the Bu/Cy group were observed after a median follow-up of 44 and 15 months, respectively. The estimated 3 year OS and DFS was 72% (95% CI: 55-98%) and 58% (95% CI: 39-77%) in the TBI and 70% (95% CI: 51-89%) for DFS and OS in the Bu/Cy group. We conclude that the anti-leukemic effect of the Bu/Cy regimen seems to be at least as effective as the TBI/Cy combination in unrelated stem cell transplantation of CML patients, with no graft failures, but that it correlates with a higher incidence of liver toxicity, hemorrhagic cystitis and chronic GVHD. Longer follow-up is necessary to determine the late relapse rate and late toxicity.

A fludarabine-based dose-reduced conditioning regimen followed by allogeneic stem cell transplantation from related or unrelated donors in patients with myelodysplastic syndrome.

We investigated the feasibility and efficacy of a fludarabine-based dose-reduced conditioning regimen followed by stem cell transplantation from related (n = 5) or unrelated HLA-matched donors (n = 7) in 12 patients with high risk MDS, who were not eligible for a standard myeloablative conditioning regimen. The conditioning regimen consisted of fludarabine 30 mg/m(2) daily for 6 days, busulfan 4 mg/kg daily for 2 days and anti-thymocyte globulin (ATG, rabbit) 10 mg/kg daily for 4 days in 11 patients, while one patient received fludarabine, ATG, cyclophosphamide and thiotepa. Graft-versus-host disease prophylaxis consisted of cyclosporine and a short course of methotrexate. The median age of the patients was 53 years (range 37-59). The median percentage of blasts in bone marrow aspirate at transplantation was 15% (range <5% to 35%). Diagnosis at transplant was RA (n = 1), RAEB (n = 5), RAEB-T (n = 5) and sAML (n = 1). A complex karyotype including monosomy 7 was noted in five patients. The reasons for using a dose-reduced conditioning regimen were prior autologous/syngeneic BMT (n = 4), active fungal infection (n = 2) or age/reduced performance status (n = 6). Engraftment was observed in all patients with complete donor chimerism. The incidence of acute GVHD (grade II-IV) was 33%. Eight patients died during follow-up due to relapse (n = 4), liver toxicity (n = 2), aspergillus (n = 1) or aGVHD grade IV (n = 1). After a median follow-up of 19 months, the 2-year estimated disease-free survival is 12% (95% CI: 2-23%) and the overall survival is 26% (95% CI: 4-52%). Fludarabine dose-reduced conditioning prior to allogeneic stem cell transplantation in high risk MDS patients, who were not eligible for standard transplantation, resulted in stable engraftment with complete chimerism, but the toxicity and relapse rate were considerable.

Use of a five-agent GVHD prevention regimen in recipients of unrelated donor marrow.

A five-agent GVHD prophylaxis programme consisting of cyclosporin A, methotrexate, anti-thymocyte-globulin, pentaglobin and metronidazol was given to 48 recipients of unrelated donor marrow with chronic myelogenous leukemia, acute leukemia, myelodysplastic syndromes, and familiar lymphocytic hemophagocytosis of an average age of 33.5 (0.6-56) years. GVHD grades II-IV occurred in 18 patients (39%) and grades III-IV in five patients (11%). Chronic GVHD developed in nine patients (23%), three limited and six extensive. Fifteen patients died. Clinical relapse was detected in eight patients. Four patients died as a consequence of the underlying disease and subsequent treatment, 11 patients died of transplant-related causes. After a median follow-up of 19 months, the overall and disease-free survival are 67% and 62%, respectively. Survival by age is as follows: 0-19 years: 12/13 patients; 20-39 years: 14/25 patients; 40-59 years: 7/10 patients. The five-agent GVHD prophylaxis regimen is effective. Matched-unrelated donor transplants can be carried out safely in patients younger than 50 years of age. The results in patients younger than 20 years of age should encourage matched-unrelated donor transplants at earlier stages of the disease.

In vivo T cell depletion with pretransplant anti-thymocyte globulin reduces graft-versus-host disease without increasing relapse in good risk myeloid leukemia patients after stem cell transplantation from matched related donors.

One-hundred and two patients with good risk myeloid leukemia (CML first chronic phase or AML first CR) were transplanted from HLA-related donors after conditioning with (n = 45) or without anti-thymocyte globulin (ATG) (n = 57). One graft failure was observed in the non-ATG and none in the ATG group. The median time to leukocyte engraftment (> 1 x 10(9)/l) was 16 (range 12-33) in the ATG group and 17 days (range 11-29) in the non-ATG group (NS) and for platelet engraftment (> 20 x 10(9)/l) 24 and 19 days (P = 0.002), respectively. Acute GVHD grade II-IV was observed in 47% of the non-ATG and in 20% of the ATG group (P = 0.004). Grade III/IV GVHD occurred in 7% of the ATG and in 32% of the non-ATG group (P = 0.002). Chronic GVHD was seen in 36% and 67% (P = 0.005), respectively. After a median follow-up of 48 months (range 2-128), the 5-year estimated OS is 66% (95% KI: 51-81%) for the ATG group and 59% (95% KI: 46-72%) for the non-ATG group (NS). The 5-year estimated DFS is 64% (95% KI: 50-78%) for ATG and 55% (95% KI: 43-67%) for the non-ATG regimen (NS). The 5-year probability of relapse was 5% in the ATG and 15% in the non-ATG group (NS). ATG as part of the conditioning regimen leads to a significant reduction in GVHD without increase of relapse in patients with myeloid leukemia after stem cell transplantation from HLA-related donors.

Dose-dependent effect of etoposide in combination with busulfan plus cyclophosphamide as conditioning for stem cell transplantation in patients with acute myeloid leukemia.

To evaluate the efficacy and toxicity of two different etoposide (VP-16) dosages (30 or 45 mg/kg) in combination with busulfan/cyclophosphamide as conditioning therapy followed by stem cell transplantation in acute myeloid leukemia (AML), 90 patients with AML received either 30 mg/kg (n = 60) or 45 mg/kg (n = 30) etoposide in combination with busulfan (16 mg/kg) and cyclophosphamide (120 mg/kg). The stem cell source was allogeneic related bone marrow (BM) (n = 53), allogeneic unrelated BM (n = 5), allogeneic unrelated peripheral blood (PBSC) (n = 2), syngeneic BM (n = 2), autologous BM purged (n = 9) or unpurged (n = 9), autologous PBSC (n = 10). Fifty-six patients (62%) were in first CR, 26 (29%) were > first CR, and eight (9%) were transplanted in relapse. Principal toxicities in both groups were mucositis and hepatotoxicity. Forty-five mg/kg etoposide resulted in greater hepatic toxicity (P = 0.03), and a higher incidence of VOD (23 vs 12%, P = 0.04) and acute GVHD grade III/IV (13 vs 5%, NS). The treatment-related mortality was 17% in the 30 mg/kg group and 33% in the 45 mg/kg group, mainly due to infections, intestinal pneumonia and GVHD. Hematological recovery of leukocytes 1/nl was comparable in both groups (17 vs 16 days). After a median follow-up of 16 months 19% in the 30 mg/kg group and 23% in the 45 mg/kg group relapsed. In patients who had undergone allogeneic related bone marrow transplantation in first CR no relapses occurred after a median follow-up of 3 years. For all patients the 3-year estimated disease-free survival was 62% in the 30 mg/kg group and 40% in the 45 mg/kg group (P = 0.03). For patients in first CR who underwent allogeneic related stem cell transplantation the 3 year disease-free survivals were 80% and 66%, respectively (P = 0.4). We conclude that etoposide 30 mg/kg or 45 mg/kg in combination with busulfan/cyclophosphamide is a highly active regimen for bone marrow transplantation of patients with AML with a low relapse rate. However, conditioning with 30 mg/kg rather than 45 mg/kg etoposide resulted in less toxicity and a better overall survival due to a lower transplant-related mortality. Bone Marrow Transplantation (2000) 26, 711-716.

Intensified conditioning regimen in bone marrow transplantation for Philadelphia chromosome-positive acute lymphoblastic leukemia.

We investigated an intensified conditioning regimen including fractionated total body irradiation (12 Gy), etoposide (30-45 mg/kg) and cyclophosphamide (120 mg/kg), followed by autologous (n = 5), allo-related (n = 13) or allo-unrelated (n = 6) bone marrow (n = 22) or peripheral stem cell (n = 2) transplantation in patients with Philadelphia chromosome-positive acute lymphoblastic leukemia. One patient received busulfan (16 mg/kg) instead of TBI. Nineteen patients were transplanted in 1CR, two in 2CR, one in 1PR and two in relapse. Major toxicity was mucositis grade II according to the Bearman scale in all patients. The treatment-related mortality was 25%, mainly due to infection or GVHD after allogeneic transplantation. After a median follow-up of 45 months (range 2-93), nine patients (37.5%) remain alive in CR. Nine patients (37.5%) relapsed and eight (33.3%) of these subsequently died. After autologous transplantation, four of five patients (80%) relapsed and died. Late relapse was seen after allogeneic, as well as autologous transplantation, at 33 and 59 months, respectively. The Kaplan-Meier estimate of leukemia-free survival for all patients is 38% at 3 years (95% CI: 18-58%) and 35% at 5 years (95% CI: 15-55%). For allogeneic transplants in first CR (n = 15) the estimate of disease-free survival was 46% at 3 years (95% CI: 19-73%) and 34% at 5 years (95% CI: 17-51%). Patients aged below 30 years had a better estimated overall survival at 3 years (61% vs 11%, P < 0.001). The bcr-abl fusion transcript (p210 vs p190 vs p210/190) did not affect disease-free or overall survival. In our experience, an intensified conditioning regimen seems to improve the results of bone marrow transplantation in patients with Ph+ acute lymphoblastic leukemia. However, the high relapse rate warrants novel approaches to enhance anti-leukemic efficacy.

Interference of cytokeratin-20 and mammaglobin-reverse-transcriptase polymerase chain assays designed for the detection of disseminated cancer cells.

Several reverse-transcriptase polymerase chain reaction (rtPCR) assays have been designed for the detection of disseminated cancer cells. The specificity of these discussed molecular approaches is controversial. Biological interference of the cytokeratin-20 and mammaglobin rtPCR assays has been investigated. Cell lines of different lineages and bone marrow and peripheral stem cells from patients without epithelial cancer have been examined for the transcription of the cytokeratin-20 (CK20) and mammaglobin messages prior to and after stimulation with different cytokines in a total of 370 liquid cultures. Amplification of both messages from clinical samples prior to stimulation does not support the high specificity for the detection of disseminated epithelial cancer cells as reported. Cytokeratin-20 was amplified from the chronic myeloic leukemia (CML)-derived line K562. Transcription was not influenced by cytokines, either in cell-line experiments or in clinical samples. The thesis of a low-level background transcription in granulocytes is supported. Mammaglobin was induced in cell lines significantly by GM-CSF and in clinical samples additionally by several more cytokines. These results indicate that under certain conditions involving cytokine production, the use of mammaglobin rtPCR for the detection of epithelial cancer cells could be limited. In conclusion, the mechanism of interference of both rtPCR assays are completely different and further research is necessary before the cytokeratin-20 or mammaglobin rtPCR could become standard methods for the detection of disseminated epithelial cancer cells. These factors leading to so-called false-positive results have to be considered in future applications of rtPCR for the detection of minimal residual disease.

CD34(+)-selected stem cell boost for delayed or insufficient engraftment after allogeneic stem cell transplantation.

BACKGROUND: Poor graft function without rejection may occur after stem cell transplantation (SCT). CD34(+) stem cell boost (SCB) can restore marrow function but may induce or exacerbate GvHD. We therefore investigated the feasibility and efficacy of CD34(+)-selected SCB in some patients with poor graft function. We present the results for eight patients (median age 46 years) transplanted initially for myelofibrosis, acute leukemia, myeloma and NHL. Six patients had received HLA-matched and two mismatched grafts (PB, BM; n=5, 3). After a median of 128 days post-transplant, the median leukocyte and platelet counts were, respectively, 2.05/nL and 18/nL. None had achieved platelet counts >50/nL even though donor chimerism was >95% in seven recipients. METHODS: Positive selection of CD34(+) stem cells was performed on a CliniMACS device, observing GMP and achieving a median of 98.5% purity. The patients received a median of 1.7 x 10(6)/kg CD34(+) cells and 2.5 x 10(3)/kg CD3(+) T lymphocytes. RESULTS: Hemograms at days +30, +60 and +90, respectively, showed steadily increasing median leukocyte (2.55, 3.15 and 4.20/nL) and platelet (29, 39 and 95/nL) counts. After a median follow-up of 144 days, five patients remained alive. No patient had developed acute or chronic GvHD. One patient died of leukemic relapse and two others of systemic mycosis. DISCUSSION: These preliminary results point to the possibility of safely improving graft function using CD34(+) positively selected stem cells without necessarily increasing the incidence of GvHD in patients with poor graft function post-SCT. Experience with more patients and longer follow-up should clarify the optimal role for this procedure.

Counselling for prenatal diagnosis of sickle cell disease and beta thalassaemia major: a four year experience.

A non-directive programme of prenatal counselling was used during a four year period. Forty-three couples at risk for having a baby with a haemoglobinopathy were identified. Prenatal diagnosis was offered in 19 pregnancies to 14 couples at risk of having a baby with sickle cell anaemia and in two pregnancies in two couples at risk of a baby with beta thalassaemia major, who presented before the 18th week of pregnancy. Six couples at risk for sickle cell anaemia accepted prenatal diagnosis in 10 pregnancies, as did both couples at risk for thalassaemia. Couples who were eligible for prenatal diagnosis but refused it tended not to have been informed about sickle cell disease before counselling, one partner was more frequently absent at the time of the initial counselling session, or they either had no children with sickle cell disease or the children were not severely affected. Other factors influencing their decision included a poor obstetric history and rejection of abortion, mainly on moral grounds. The approximately 50% uptake of prenatal diagnosis in this initial study highlights the complex issues involved. Our experience indicates that with systematic screening and counselling in the antenatal clinic, and with increased awareness of the haemoglobinopathies, couples at risk will be in a better position to make informed decisions.

Genetic relationship between the 3'-VNTR and diallelic apolipoprotein B gene polymorphisms: haplotype analysis in individuals of European and south Asian origin.

The genetic relationship amongst apolipoprotein B (apo B) gene polymorphisms (signal peptide insertion/deletion (Leu-Ala-Leu-16/-14), XbaI (Thr2488), EcoRI (Glu4154----Lys), Asn4311----Ser, 3'-VNTR) has been investigated in samples of South Asian (Indian) and Swedish individuals. The frequency distribution of alleles at all these sites was found to be significantly different between the South Asian and the Swedish samples (deletion allele: 0.20 v. 0.31, X+ (presence of XbaI cutting site): 0.29 v. 0.55, R- (absence of EcoRI cutting site): 0.11 v. 0.19, Ser4311: 0.45 v. 0.19). The distribution of allele frequencies at the VNTR site was bimodal in both populations. However, in South Asians, the most common allele was a 35 repeat unit allele, whilst in the Swedish sample, and in all other reports from Caucasian samples the 37 repeat unit allele was the most frequent (South Asian v. Swedish: 35 allele: 0.36 v. 0.19, 37 allele: 0.25 v. 0.48). Furthermore, four new alleles at the apo B gene 3'-VNTR site (15, 17, 32, 38 repeat unit alleles) were observed in South Asians, of which two (15 and 17 repeat unit alleles) were well outside the bimodal distribution. In both samples, strong linkage disequilibrium and allelic association were detected between alleles at the 3'-VNTR and each of the other sites, and also between the ins/del and XbaI sites and between the XbaI and Asn4311----Ser sites. The same five common haplotypes as defined by ins/del, XbaI, EcoRI and Asn4311----Ser were found to be present in both samples comprising 97 and 99% of the haplotypes observed in the South Asian and Swedish samples respectively. Detailed analysis revealed the predominant occurrence of certain 3'-VNTR alleles on specific haplotypes and demonstrates the usefulness of the VNTR site for haplotyping and representative association studies. A model for the evolutionary relationship of the major haplotypes including the 3'-VNTR site is presented. These findings support a mechanism of replication slippage as a major factor for the generation of new alleles at the apo B 3'-VNTR locus rather than unequal crossing over.

Allogeneic bone marrow transplantation for refractory mantle cell lymphoma.

We report about a 28-year-old woman with relapsed mantle cell lymphoma (MCL, centrocytic lymphoma according to the Kiel classification) refractory to salvage chemotherapy. The patient underwent allogeneic bone marrow transplantation from a HLA-identical brother after myeloablative chemotherapy consisting of busulfan, etoposide, and cyclophosphamide. The patient experienced hepatic toxicity (grade I), mucositis (grade II) according the Bearman scale, and graft versus host disease of the skin (grade II) and showed stable engraftment with complete chimerism on day 15 after bone marrow transplantation. Eight years after transplantation, the patient is still disease free and in good condition without any late side effects. This report suggests a curative potential of allogeneic stem cell transplantation in MCL.

Donor lymphocyte infusion enhances remission status in patients with persistent disease after allografting for multiple myeloma.

Two patients with persistent disease after allografting for multiple myeloma received donor T-cell lymphocyte infusion (DLI) (1.5 x 10(8) and 7 x 10(7)) to induce a graft-vs.-myeloma effect for further tumour regression after withdrawal of immunosuppression. The interval between stem cell transplantation and DLI was 8 and 14 months respectively. Both patients converted from partial to complete remission, lasting 12+ and 28+ months. Immunofixation became negative after 3 and 4 months. The main toxicity was grade II and III acute graft-vs.-host disease (GvHD) and limited or extensive chronic GvHD in each patient. We conclude that DLI induced further tumour reduction in patients with persistent disease after allografting for multiple myeloma.

A randomized comparison of once versus twice daily recombinant human granulocyte colony-stimulating factor (filgrastim) for stem cell mobilization in healthy donors for allogeneic transplantation.

To evaluate the schedule dependency of granulocyte colony-stimulating factor (G-CSF) (filgrastim) for stem cell mobilization, we conducted a randomized comparison in 50 healthy donors, with one subcutaneous daily injection of 10 microg/kg G-CSF (n = 25) compared with twice injections daily of 5 microg/kg G-CSF (n = 25). The two groups were well balanced for age, body weight and sex. G-CSF application was performed on an out-patient basis and leukapheresis was started in all donors on day 5. The most frequent side-effects of G-CSF were mild to moderate bone pain (88%), mild headache (72%), mild fatigue (48-60%) and nausea (8%) without differences between the two groups. The CD34(+) cell count in the first apheresis was 5.4 x 10(6)/kg donor weight (range 2.8-13.3) in the 2 x 5 microg/kg group compared with 4.0 x 10(6)/kg (range 0.4-8.8) in the 1 x 10 microg/kg group (P = 0.007). The target of collecting > 3.0 x 10(6) CD34(+) cells/kg donor weight with one apheresis procedure was achieved in 24/25 (96%) donors in the 2 x 5 microg/kg group and in 17/25 (68%) donors in the 1 x 10 microg/kg group. The target of collecting > 5.0 x 10(6) CD34(+) cells/kg in the first apheresis was achieved in 64% in the 2 x 5 microg/kg group, but in only 36% in the 1 x 10 microg/kg group. The progenitor cell assay for granulocyte-macrophage colony-forming units (CFU-GM) and erythroid burst-forming units (BFU-E) was higher in the 2 x 5 microg/kg group than in the 1 x 10 microg/kg group (7.0 vs. 3.5 x 10(5)/kg, P = 0.01; 6.6 vs. 5.0 x 10(5)/kg; P = 0.1). Administering G-CSF (filgrastim) at a dosage of 5 microg/kg twice daily rather than 10 microg/kg once daily is recommended; this leads to a higher CD34(+) cell yield and requires fewer apheresis procedures without increasing toxicity or cost.