Thermal shock susceptibility and regrowth of Pseudomonas aeruginosa biofilms.

Biofilms on implanted medical devices cause thousands of patients each year to undergo multiple surgeries to remove and replace the implant, driving billions of dollars in increased health care costs due to the lack of viable treatment options for in situ biofilm eradication. Remotely activated localised heating is under investigation to mitigate these biofilms; however, little is known about the temperatures required to kill the biofilms. To better understand the required parameters this study investigated the thermal susceptibility of biofilms as a function of their fluidic and chemical environment during growth, as well as their propensity for regrowth following thermal shock. Pseudomonas aeruginosa biofilms were cultured in shaker plate fluidic conditions in four different growth media, then thermally shocked at various temperatures and exposure times. Biofilms were re-incubated to determine their regrowth potential following thermal shocks of various intensities. Results indicate that growth media has little impact on thermal susceptibility, while fluidic conditions strongly influence susceptibility to modest thermal shocks. This effect disappears, however, with increasingly aggressive shocks, reducing biofilm populations by up to 5 orders of magnitude. Regrowth studies indicate a critical post-shock bacterial loading (∼103 CFU/cm2) below which the biofilms were no longer viable, while biofilms above that loading slowly regrew to their previous population density.

Incorporating hydrology into climate suitability models changes projections of malaria transmission in Africa.

Continental-scale models of malaria climate suitability typically couple well-established temperature-response models with basic estimates of vector habitat availability using rainfall as a proxy. Here we show that across continental Africa, the estimated geographic range of climatic suitability for malaria transmission is more sensitive to the precipitation threshold than the thermal response curve applied. To address this problem we use downscaled daily climate predictions from seven GCMs to run a continental-scale hydrological model for a process-based representation of mosquito breeding habitat availability. A more complex pattern of malaria suitability emerges as water is routed through drainage networks and river corridors serve as year-round transmission foci. The estimated hydro-climatically suitable area for stable malaria transmission is smaller than previous models suggest and shows only a very small increase in state-of-the-art future climate scenarios. However, bigger geographical shifts are observed than with most rainfall threshold models and the pattern of that shift is very different when using a hydrological model to estimate surface water availability for vector breeding.

Ultrastructural basis of biochemical effects in a series of lethal alleles in the mouse. Neonatal and developmental studies.

The fine structure of newborn and fetal mouse liver and of newborn kidney cells homozygous for any of three albino alleles known to have multiple biochemical effects was investigated. Electron microscope studies of mutant cells revealed dilation and vesiculation of the rough endoplasmic reticulum in parenchymal liver cells, as well as dilation and other anomalies of the Golgi apparatus. These abnormalities were observed in all newborn mutants but never in littermate controls. Although they were most pronounced in liver parenchymal cells, they were found also to a lesser degree in kidney cells, but they were absent altogether in other cell types of the mutant newborn. Homozygous fetuses showed similar anomalies in the liver at 19 days of gestational age. In one of the alleles studied, mutant liver parenchymal cells were found to be abnormal as early as the 18th day of gestation. There appears to be a striking parallelism between the biochemical defects and those of the cellular membranes in homozygous mutant newborn and fetuses. Although the specific nature of the mutational effect on membrane structure remains unknown, the results are compatible with the assumption that a mutationally caused defect in a membrane component interferes with a mechanism vital in the integration of morphological and biochemical differentiation.

Transplantation of leukemic bone marrow treated with cytotoxic antileukemic antibodies and complement.

The ability of antiserum against murine L1210 leukemia to remove residual leukemia cells from murine bone marrow was investigated. Leukemic marrow was treated in vitro with antiserum and complement and used to hematologically reconstitute mice that had been irradiated with doses lethal to bone marrow. Following infusion of treated leukemic marrow, normal marrow returned without evidence of leukemia. More than 90 percent of the animals have survived for 11 months without untoward effects, suggesting that the technique may be of use in the treatment of acute leukemia in humans.

In vitro analysis of donor bone marrow following monoclonal antibody treatment for the prevention of acute graft versus host disease.

Graft-versus-host disease (GVHD) remains a major cause of morbidity and mortality following bone marrow transplantation. The in vitro removal of the GVHD-causing T-lymphocytes from donor marrow is one approach which could control this complication. Treatment of the donor bone marrow with lectins and erythrocyte-forming rosette depletion, anti-T-cell antisera or monoclonal antibodies are methods currently being tested to accomplish this. CT-2 is an immunoglobulin monoclonal antibody specific for the T-cell erythrocyte-forming rosette receptor. Bone marrow from 23 consecutive donors was treated in vitro with CT-2 and complement, prior to infusion, as a potential means of controlling GVHD. Surface marker analysis using erythrocyte-forming rosetting, and OKT-3 and OKT-11 monoclonal antibodies on paired samples of treated and untreated marrow demonstrated a mean depletion to 1% of the original number of T-cells. Proliferative responses to alloantigens and mitogens as well as cytotoxic and natural killer cell function were tested and found to be markedly reduced. Despite these effects on T-lymphocytes, viable hematopoietic stem cell colonies were retained. Clinical results following the in vitro T-lymphocyte depletion of donor bone marrow for the 8 histocompatible and 15 nonhistocompatible bone marrow transplantation are reported. Prompt engraftment with minimal GVHD, despite no posttransplant GVHD prophylaxis, was seen in seven of the matched patients. In the nonhistocompatible bone marrow transplantation, failure of engraftment occurred in 11 patients. Grades III-IV GVHD were seen in two of the four patients that engrafted despite good T-lymphocyte depletion. No predictive correlation could be found between the in vitro analysis of marrow following CT-2 treatment and clinical outcome.

Osteonecrosis as a complication of treating acute lymphoblastic leukemia in children: a report from the Children's Cancer Group.

PURPOSE: To determine the incidence, risk factors, and morbidity for osteonecrosis (ON) in children with acute lymphoblastic leukemia (ALL) treated with intensive chemotherapy including multiple, prolonged courses of corticosteroid. PATIENTS AND METHODS: The occurrence of symptomatic ON was investigated retrospectively in 1, 409 children ages 1 to 20 years old receiving therapy for high-risk ALL on Children's Cancer Group (CCG) protocol CCG-1882. RESULTS: ON was diagnosed in 111 patients (9.3% +/- 0.9%, 3-year life-table incidence). The incidence was higher for older children (> or = 10 years: 14.2% +/- 1.3% v < 10 years: 0.9% +/- 0.4%; P: <.0001), especially females 10 to 15 years old and males 16 to 20 years old (19.2% +/- 2.3% and 20.7% +/- 4.7%, respectively). In patients 10 to 20 years old, the incidence of ON was higher for females versus males (17.4% +/- 2.1% v 11.7% +/- 1.6%, respectively; P: =.03) and for patients randomized to receive two 21-day dexamethasone courses versus one course (23.2% +/- 4.8% v 16.4% +/- 4.3%, respectively; P: =.27). Among ethnic groups, whites had the highest incidence and blacks the lowest, with other groups intermediate (16.7% +/- 1.4% v 3.3% +/- 2.3% v 6.7% +/- 2.2%, respectively; P: =.003). There was no difference in event-free survival in patients with or without ON. ON was diagnosed within 3 years of starting ALL therapy in all but one patient, involved weight-bearing joint(s) in 94% of patients, and was multifocal in 74% of patients. Symptoms of pain and/or immobility were chronic in 84% of patients, with 24% having undergone an orthopedic procedure and an additional 15% considered candidates for surgery in the future. CONCLUSION: Children ages 10 to 20 years who receive intensive ALL therapy, including multiple courses of corticosteroid, are at significant risk for developing ON.

Augmented Berlin-Frankfurt-Munster therapy abrogates the adverse prognostic significance of slow early response to induction chemotherapy for children and adolescents with acute lymphoblastic leukemia and unfavorable presenting features: a report from the Children's Cancer Group.

PURPOSE: Compared with previous Children's Cancer Group (CCG) acute lymphoblastic leukemia (ALL) trials, therapy based on the Berlin-Frankfurt-Munster (BFM) 76 trial has effected an improvement in event-free survival (EFS). In an attempt to improve EFS further, CCG investigators formulated an augmented BFM (A-BFM) regimen that provides prolonged, intensified postinduction chemotherapy relative to the CCG-modified BFM regimen. PATIENTS AND METHODS: We tested A-BFM in 101 patients with ALL and unfavorable presenting features that showed slow early response (SER) to induction therapy who attained remission on day 28. Their outcome was compared with that of 251 concurrent patients with unfavorable presenting features, a rapid early response to therapy (RER), and remission by day 28, treated with CCG-BFM with or without cranial radiation (CRT). RESULTS: The 4-year EFS rate from the end of induction for SER patients treated with A-BFM was 70.8% +/- 4.6%. Seventeen patients remain in continuous remission beyond 5 years. Vincristine (VCR) neurotoxicity developed in 50% of patients, but was rarely debilitating. Allergies to Escherichia coli L-asparaginase (L-ASP) occurred in 35% of patients. Avascular necrosis of bone (AVN) developed in 9% of patients. In comparison, a concurrent RER group treated with standard BFM +/- CRT had a 4-year EFS rate of 73.1% +/- 4.6%. CONCLUSION: The toxicity of A-BFM is significant, but acceptable. Compared with historical control SER patients treated with CCG-modified BFM, A-BFM therapy appears to produce a significant improvement in EFS. This is the first study to show that intensive chemotherapy, as given in the A-BFM regimen, can abrogate the adverse prognostic significance of SER.

Response of children with high-risk acute lymphoblastic leukemia treated with and without cranial irradiation: a report from the Children's Cancer Group.

PURPOSE: Intensified intrathecal (i.t.) chemotherapy without cranial radiation therapy (CRT) prevents CNS relapse in children with low-risk and intermediate-risk acute lymphoblastic leukemia (ALL). In the current study, high-risk ALL patients who achieved a rapid early response (RER) to induction chemotherapy were randomized to receive intensive systemic chemotherapy and presymptomatic CNS therapy that consisted of either i.t. methotrexate (MTX) and CRT or intensified i.t. MTX alone. PATIENTS AND METHODS: Children (n = 636) with high-risk ALL (aged 1 to 9 years and WBC count > or = 50,000/microL or age > or = 10 years, excluding those with lymphomatous features) who achieved an RER (< or = 25% marrow blasts on day 7) to induction therapy and lacked CNS disease at diagnosis were randomized to receive systemic therapy with either i.t. MTX and CRT (regimen A, n = 317) or intensified i.t. MTX alone (regimen B, n = 319). RESULTS: Interim analysis in July 1993 revealed 3-year event-free survival (EFS) estimates of 82.1% +/- 4.0% (SD)and 70.4% +/- 4.2% for patients treated on regimens A and B, respectively (P = .004). As of January 1996, outcome had changed: 5-year EFS estimates were 69.1% +/- 3.4% and 75.0% +/- 2.7% for regimens A and B, respectively (P = 0.50). Marrow relapses comprised 57 events on regimen A and 43 events on regimen B. Fewer late events occurred on regimen B. CONCLUSION: For high-risk pediatric ALL patients who show an RER to induction therapy and are treated with systemic Children's Cancer Group (CCG)-modified Berlin-Frankfurt-Munster (BFM) chemotherapy, presymptomatic CNS therapy that consists of either i.t. MTX plus CRT or intensified i.t. MTX alone results in a similar 5-year EFS outcome. Furthermore, intensified i.t. MTX may protect against late bone marrow relapse.

Clinical features and treatment outcome of childhood T-lineage acute lymphoblastic leukemia according to the apparent maturational stage of T-lineage leukemic blasts: a Children's Cancer Group study.

PURPOSE: Leukemic cells from T-lineage acute lymphoblastic leukemia (ALL) patients are thought to originate from T-lymphocyte precursors corresponding to discrete stages of T-cell ontogeny. Here we sought to determine the influence of leukemic cell apparent maturational stage on treatment outcomes in pediatric T-lineage ALL. PATIENTS AND METHODS: From 1983 through 1993, 407 pediatric T-lineage ALL patients were enrolled onto two sequential series of risk-adjusted treatment protocols of the Children's Cancer Group. In the current analysis, T-lineage ALL patients were immunophenotypically classified as follows: CD7+ CD2- CD5- pro-thymocyte leukemia (pro-TL), CD7+ (CD2 or CD5)+ CD3- immature TL, and CD7+ CD2+ CD5+ CD3+ mature TL. RESULTS: Similar induction outcomes of 91.4%, 97.1%, and 98.3% were obtained by the pro-, immature, and mature TL groups, respectively. Four-year event-free survival (EFS) was lower for pro-TL patients (57.1%; SD = 8.4%,) compared with immature and mature TL patients (68.5%; SD = 3.5%; and 77.1%; SD = 4.0%, respectively) with an overall significance of .05 (log-rank test) or .04 (log-rank trend test). Relative hazards rates (RHR) were 2.11 and 1.22 for pro-TL and immature TL versus mature TL, respectively. Highly significant differences were found for overall survival (P = .005, log-rank test; P = .009, log-rank trend test). Multivariate analysis confirmed that the prognostic influence of ontogeny grouping was independent of that of other prognostic factors. CONCLUSION: Leukemic cells of the pro-TL maturation stage identify a small subgroup of T-lineage ALL patients who have a significantly worse EFS outcome than patients whose cells are of a more mature stage of development.

Cytoreduction and prognosis in acute lymphoblastic leukemia--the importance of early marrow response: report from the Childrens Cancer Group.

PURPOSE: To quantify the residual marrow lymphoblast fraction that best defines patients at high risk for relapse, and the optimal time for assessment during remission induction. PATIENTS AND METHODS: The residual lymphoblast percentage was evaluated on day 7 (n = 220) and day 14 (n = 205) during a four- or five-drug induction in patients with poor prognostic factors. The rate of cytoreduction was related to event-free survival (EFS) and other factors. RESULTS: On the New York (NY) regimen, 68%, 14%, and 18%, and on the Berlin-Frankfurt-Munster (BFM) regimen, 56%, 15%, and 29% of patients had M1 (< 5% blasts), M2 (5-25%), or M3 (> 25%) responses on day 7 (P = .075). On day 14, the corresponding values were 87%, 6%, 7% on NY and 84%, 8%, 8% on BFM. For patients who achieved remission by day 28 and a day-7 marrow rating of M1, M2, or M3, the 6-year EFS rate was 78%, 61%, and 49% (P < .001). The day-14 ratings predicted for a 72%, 32%, or 40% EFS (P < .001). Patients with 5% to 10% blasts day 7 had three times as many events as those with less than 5% and had no better EFS than those with 11% to 25% blasts. Patients with a WBC count more than 200,000/microL at diagnosis and an M1 day 7 marrow had an EFS rate of 69%, while for those with M2 or M3, the EFS rate was 41%. Day-7 marrow had greater prognostic significance than the day-14 evaluation. For slow responders on day 7, the day-14 marrow provided additional information. EFS for patients who achieved M1 by day 14 was 65%. EFS decreased to 20% for those still M2 or M3 on day 14. Day-7 and -14 evaluations had significance for patients of all ages and WBC levels. CONCLUSION: Marrow aspiration on day 7 of therapy provided more useful information than that on day 14. However, day-14 marrow provided additional information for patients with a poor day-7 response. The rate of cytoreduction is a powerful, independent prognostic factor that can identify patients with a slow early response who are at risk for a short remission duration.

MOPP or radiation in addition to ABVD in the treatment of pathologically staged advanced Hodgkin's disease in children: results of the Children's Cancer Group Phase III Trial.

PURPOSE: A randomized trial designed to compare mechlorethamine, vincristine, procarbazine, and prednisone (MOPP)/doxorubicin, bleomycin, vinblastine, and daccarbazine (ABVD) (regimen A) with ABVD plus low-dose regional (extended-field) radiation therapy (EF RT) (regimen B) for the treatment of children and adolescents with stages III and IV Hodgkin's disease was conducted by the Children's Cancer Group (CCG-521) from 1986 until 1990. PATIENTS AND METHODS: One hundred eleven eligible patients were randomized, 57 to regimen A and 54 to regimen B. All patients had pathologically verified stage III or stage IV Hodgkin's disease. RESULTS: Overall survival (S) is 87% at 4 years and event-free survival (EFS) is 82%. Patients randomized to ABVD plus EF RT have a 4-year EFS of 87% compared with 77% for patients randomized to MOPP/ABVD (P = .09, two-sided). Patients randomized to ABVD plus EF RT have a 4-year S of 90% compared with 84% for patients randomized to MOPP/ABVD (P = .45, two-sided). Significant prognostic factors in multivariate analysis for EFS are stage of disease, erythrocyte sedimentation rate (ESR) at diagnosis, liver size at diagnosis, and, among stage III patients, the size of the mediastinal mass at diagnosis. The acute toxicities of treatment are largely hematopoietic in nature, whereas acute pulmonary and cardiac toxicities are modest and not limiting. CONCLUSION: The results of this study show that, in advanced-stage Hodgkin's disease in children, equivalent results can be obtained by the addition of either MOPP or low-dose EF RT to the ABVD regimen; whether the addition of either contributes to outcome was not addressed in this study and will require additional testing. It is clear, however, that MOPP chemotherapy can safely be eliminated from front-line combination chemotherapy regimens for advanced Hodgkin's disease in pediatric patients.

Uniform approach to risk classification and treatment assignment for children with acute lymphoblastic leukemia.

PURPOSE: To define more uniform criteria for risk-based treatment assignment for children with acute lymphoblastic leukemia (ALL), the Cancer Therapy Evaluation Program (CTEP) of the National Cancer Institute (NCI) sponsored a workshop in September 1993. Participants included representatives from the Childrens Cancer Group (CCG), Pediatric Oncology Group (POG), Dana-Farber Cancer Institute (DFCI), St Jude Children's Research Hospital (SJCRH), and the CTEP. METHODS: Workshop participants presented and reviewed data from ALL clinical trials, using weighted averages to combine outcome data from different groups. RESULTS: For patients with B-precursor (ie, non-T, non-B) ALL, the standard-risk category (4-year event-free survival [EFS] rate, approximately 80%) will include patients 1 to 9 years of age with a WBC count at diagnosis less than 50,000/microL. The remaining patients will be classified as having high-risk ALL (4-year EFS rate, approximately 65%). For patients with T-cell ALL, different treatment strategies have yielded different conclusions concerning the prognostic significance of T-cell immunophenotype. Therefore, some groups/institutions will classify patients with T-cell ALL as high risk, while others will assign risk for patients with T-cell ALL based on the uniform age/WBC count criteria. Workshop participants agreed that the risk category of a patient may be modified by prognostic factors in addition to age and WBC count criteria, and that a common set of prognostic factors should be uniformly obtained, including DNA index (DI), cytogenetics, early response to treatment (eg, day-14 bone marrow), immunophenotype, and CNS status. CONCLUSIONS: The more uniform approach to risk-based treatment assignment and to collection of specific prognostic factors should increase the efficiency of future ALL clinical research.

Frequency and clinical significance of cytogenetic abnormalities in pediatric T-lineage acute lymphoblastic leukemia: a report from the Children's Cancer Group.

PURPOSE: Nonrandom chromosomal translocations are frequently observed in pediatric patients with acute lymphoblastic leukemia (ALL). Specific translocations, such as t(4;11) and t(9;22), identify subgroups of B-lineage ALL patients who have an increased risk of treatment failure. The current study was conducted to determine the prognostic significance of chromosomal translocations in T-lineage ALL patients. MATERIALS AND METHODS: The study included 169 children with newly diagnosed T-lineage ALL enrolled between 1988 and 1995 on risk-adjusted protocols of the Children's Cancer Group (CCG) who had centrally reviewed cytogenetics data. Outcome analyses used standard life-table methods. RESULTS: Presenting features for the current cohort were similar to those of concurrently enrolled patients for whom cytogenetic data were not accepted on central review. The majority of patients (80.5%) were assigned to CCG protocols for high-risk ALL and 86.4% had pseudodiploid (n = 80) or normal diploid (n = 66) karyotypes; modal chromosome number was not a significant prognostic factor. Overall, 103 of 169 (61%) patients had an abnormal karyotype, including 31 with del(6q), 29 with 14q11 breakpoints, 15 with del(9p), 11 with trisomy 8, nine with 11q23 breakpoints, nine with 14q32 translocations, and eight with 7q32-q36 breakpoints. Thirteen patients had the specific 14q11 translocation t(11;14)(p13;q11) and all were classified as poor risk. Patients with any of these translocations had outcomes similar to those with normal diploid karyotypes. CONCLUSION: Chromosomal abnormalities, including specific nonrandom translocations, were frequently observed in a large group of children with T-lineage ALL, but were not significant prognostic factors for this cohort. Thus, contemporary intensive treatment programs result in favorable outcomes for the majority of T-lineage ALL patients, regardless of karyotypic abnormalities, and such features do not identify patients at higher risk for relapse.

Treatment outcome and prognostic factors for infants with acute lymphoblastic leukemia treated on two consecutive trials of the Children's Cancer Group.

PURPOSE: Infants represent a very poor risk group for acute lymphoblastic leukemia (ALL). We report treatment outcome for such patients treated with intensive therapy on consecutive Children's Cancer Group (CCG) protocols. PATIENTS AND METHODS: Between 1984 and 1993, infants with newly diagnosed ALL were enrolled onto CCG-107 (n = 99) and CCG-1883 (n = 135) protocols. Postconsolidation therapy was more intensive on CCG-1883. On both studies, prophylactic treatment of the CNS included both high-dose systemic chemotherapy and intrathecal therapy, in contrast to whole-brain radiotherapy, which was used in earlier studies. RESULTS: Most patients (>95%) achieved remission with induction therapy. The most frequent event was a marrow relapse (46 patients on CCG-107 and 66 patients on CCG- 1883). Four-year event-free survival was 33% (SE = 4.7%) on CCG-107 and 39% (SE = 4.2%) on CCG- 1883. Both studies represent an improvement compared with a 22% (SE = 5.1%) event-free survival for historical controls. Four-year cumulative probabilities of any marrow relapse or an isolated CNS relapse were, respectively, 49% (SE = 5%) and 9% (SE = 3%) on CCG-107 and 50% (SE = 5%) and 3% (SE = 2%) on CCG-1883, compared with 63% (SE = 6%) and 5% (SE = 3%) for the historical controls. Independent adverse prognostic factors were age less than 3 months, WBC count of more than 50,000/microL, CD10 negativity, slow response to induction therapy, and presence of the translocation t(4;11). CONCLUSION: Outcome for infants on CCG-107 and CCG- 1883 improved, compared with historical controls. Marrow relapse remains the primary mode of failure. Isolated CNS relapse rates are low, indicating that intrathecal chemotherapy combined with very-high-dose systemic therapy provides adequate protection of the CNS. The overall unsatisfactory outcome observed for the infant ALL population warrants the future use of novel alternative therapies.

Children's Cancer Group trials in childhood acute lymphoblastic leukemia: 1983-1995.

Since 1968, the Children's Cancer Group (CCG) has treated more than 16,000 children with acute lymphoblastic leukemia (ALL). Herein, we report improvements obtained in CCG trials during two successive series of studies (1983-1988 and 1989-1995). Overall, 10-year EFS was 62% +/- 10% for the 1983-1988 series and 72% +/- 1% for the 1988-1995 series (P< 0.0001). Five-year cumulative rates of isolated CNS relapses were 5.9% and 4.4%. Therapy based on the Berlin-Frankfurt-Münster 76/79 study improved outcomes for intermediate and higher risk patients in the first series. For intermediate risk patients, delayed intensification (DI) was most crucial for improved outcome and cranial irradiation was safely replaced with maintenance intrathecal methotrexate, providing patients received intensified systemic therapy. In the second series, randomized trials showed better outcome with one vs no DI phase for lower risk patients, with two vs one DI phase for intermediate risk patients, and with the CCG 'augmented regimen' for higher risk patients with a slow day 7 marrow response. Cranial irradiation was safely replaced with additional intrathecal methotrexate for higher risk patients with a rapid day 7 marrow response. In a subsequent study, substitution of dexamethasone in place of prednisone in induction and maintenance improved outcome for standard risk patients. All patients received dexamethasone in DI. These successful treatment strategies form the basis for our current ALL trials.

Cytogenetic studies of infant acute lymphoblastic leukemia: poor prognosis of infants with t(4;11) - a report of the Children's Cancer Group.

Infants less than 1 year of age at diagnosis of acute lymphoblastic leukemia (ALL) have a poor prognosis, which has been attributed primarily to a breakpoint in chromosomal band 11q23 or the MLL gene. Most infants with an 11q23 breakpoint have a t(4;11)(q21 ;q23). We studied the cytogenetics of the leukemia cells of 56 infants on CCG-1883, a single-arm clinical treatment protocol for infant ALL. Twenty-one patients had t(4;11)(q21;q23), seven had other rearrangements with breakpoints in 11q23 (other 11q23), 16 had normal chromosomes, two had t(1;19)(q32;p13), one had >50 chromosomes, and nine had non-recurring structural abnormalities. To determine whether there is a difference in outcome for infants with t(4;11), other 11q23 and the remaining patients, we compared event-free survival (EFS) and other clinical and laboratory features of the above infants. Infants without t(4;11) and those with other 11q23 rearrangements had significantly better EFS than those with t(4;11) (P= 0.007 and P= 0.02, respectively). t(4;11) correlated with age less than 6 months and with CD10 negativity, both of which also were poor prognostic indicators. After adjustment for age, there was still a significant difference in EFS between patients with t(4;11) and those with other 1lq23 rearrangements (P=0.02), and between patients with t(4;11) and those without t(4;11) (P=0.04). Among CD10 negative patients, t(4;11) was associated with a worse EFS (P=0.01). Multivariate analysis showed that after adjusting for a variety of clinical and laboratory features, t(4;11) was the most important prognostic factor for poor outcome, and patients with other 11q23 rearrangements had as good an outcome as the remaining patients without t(4;11).

A rapid, single step technique for determination of B and T cell surface markers in malignant lymphoid cells.

The immunological classification of lymphocytic subpopulations has become increasingly important in the treatment of hematologic malignancy. In the present study, the standard techniques for the identification of subsets of both normal and malignant lymphocytes were compared to a new one step procedure for the simultaneous assessment and identification of cells with T or B cell surface markers. With this new technique, cells with T lymphocyte markers form SRBC rosettes and cells with B lymphocyte markers form bead rosettes after incubation with microspheres coated with an anti-immunoglobulin antibody. In the present study, analysis of patient material using this simultaneous marker method produced results similar to standard techniques. In addition, the simultaneous E-rosette and immunobead method for detection of lymphocyte subpopulations permits direct examination of cell morphology, is easier to perform, and does not require fluorescence microscopy.

Relapse of host leukemic lymphoblasts following engraftment by an HLA-mismatched marrow transplant: mechanisms of escape from the "graft versus leukemia" effect.

Leukemic relapses and graft versus host disease (GvHD) remain major complications following allogeneic bone marrow transplantation for leukemia. We present clinical and laboratory details for an eight-year-old boy who received a T-cell-depleted HLA-mismatched marrow transplant as therapy for acute lymphoblastic leukemia (ALL) in second remission. Engraftment with donor marrow was prompt and without any acute GvHD. Nevertheless, the patient's original ALL recurred and proved fatal. The patient remained a chimera with persistent donor lymphocytes present at the time of posttransplant relapse and subsequent to treatment with unsuccessful reinduction chemotherapy. In vitro immune studies showed that these leukemic cells could be recognized and destroyed by the donor's lymphocytes. The relapse itself suggests, however, that the donor's lymphocytes did not effectively destroy the patient's histoincompatible ALL cells in vivo following establishment of the chimeric state. Potential mechanisms are presented to account for this presumed "escape" from the postulated "graft versus leukemia" effect.

Transplantation of HLA-haploidentical T-cell-depleted marrow for leukemia: autologous marrow recovery with specific immune sensitization to donor antigens.

Autologous marrow recovery without engraftment of donor marrow was observed after bone marrow transplantation (BMT) for two patients with acute lymphoblastic leukemia. Each had received marrow from a haploidentical mixed lymphocyte culture (MLC) reactive donor after pretransplant conditioning with total body irradiation and high-dose cyclophosphamide. To minimize graft-vs-host disease, the marrow was depleted of T cells in vitro by treatment with a monoclonal anti-T-cell antibody and complement. Two weeks after each transplant, reactive lymphocytes were noted transiently in the blood of each patient. Analysis of karyotype, HLA type, and in vitro MLC responsiveness proved the lymphocytes to be of host, not donor, origin. MLC studies showed rapid proliferative responses specifically to stimulating cells from the BMT donor, indicating in vivo sensitization to donor antigens. Return of hematopoietic function was markedly delayed, but it eventually normalized after several months, without evidence of chimerism. These studies confirm that some immune and hematopoietic stem cells of host origin survive the high-dose chemoradiotherapy used as transplant conditioning. Because these immune cells are specifically reactive to donor alloantigens, more potent suppression of host immunity may be needed to prevent nonengraftment of T-cell-depleted, HLA-mismatched bone marrow.

Depletion of T cells from bone marrow for allogeneic transplantation: method for treatment of bone marrow in bulk.

T lymphocytes were depleted from donor marrow for 23 patients undergoing allogeneic bone marrow transplantation using an anti-T-cell antibody, CT-2, and complement. The methodology is described in detail for in vitro depletion of large quantities of bone marrow. The extent of T-lymphocyte depletion using various T-cell markers, the percent of marrow lost in the processing and quantity of antibody, and complement needed are presented. These techniques for in vitro T-lymphocyte depletion were reproducible and did result in an average final yield of 47% of the harvested donor marrow.