Acute lymphoblastic leukaemia cells express CCR7 but not higher amounts of IL-10 after CD40 ligation.

OBJECTIVE: Production of cytokines that support T-cell activation and proliferation and migration to lymph nodes is one of the most important terms of cancer vaccine development. In previous studies we and others used CD40 ligation to obtain higher expression of co-stimulatory and adhesion molecules on leukaemic cells from children with acute lymphoblastic leukaemia (ALL). This time we assess the cytokine and chemokine gene expression profile in CD40-stimulated ALL cells. MATERIAL AND METHODS: Malignant cells from 25 children with BCP-ALL were stimulated (or not) with huCD40LT and rIL-4 for 96 h. Eleven different molecule, cytokine and chemokine mRNAs levels (CCR7, IL-23, TGF-beta-IP, IFN-gamma, IL-10, CD1a, CD40, CD54, CD80, CD83, CD86) were determined using the real-time PCR technique with TaqMan chemistry using ready-to-use low-density arrays for gene expression by Applied Biosystems. RESULTS: 1) Increases in mRNA levels for CD40, CD54 and CD80 after CD40L and IL-4 stimulation were observed, 2) CCR7 mRNA expression was higher after CD40 ligation than before the culture (p = 0.002), 3) IL-10 mRNA expression was higher after the culture with medium than before the culture (p = 0.01). CONCLUSIONS: The results show that leukaemia-derived dendritic cells obtained with CD40 ligation express CCR7 - chemokine is involved in migration to lymph nodes and does not produce higher amounts of IL-10, a potent immunosuppressive cytokine. Our preclinical findings could be used in the design of immunotherapy trials for the treatment of children with ALL.

Twenty years of Polish experience with three consecutive protocols for treatment of childhood acute myelogenous leukemia.

Until 1983, results of treatment of acute myelogenous leukemia (AML) in Poland with different regimens were very poor. In 1983, the Polish Pediatric Leukemia/Lymphoma Study Group introduced a unified treatment protocol--a modified version of BFM-83 protocol. This led to an increase in the curability of AML from 15% to approximately 32%. In 1994, a modification was made: the high-risk patients (>5% blasts in bone marrow on day 15 of therapy and all M5 cases) received two additional cycles with intermediate-dose cytarabine (ID-ARAC). This led to a nonsignificant improvement in the 5-year event-free survival (EFS) rate from 32 to 36%. A new treatment protocol employing idarubicin in place of daunorubicin was introduced in 1998 and produced better initial responses, increase in the number of patients attaining remission after induction therapy and proportional increase of standard-risk patients. The probability of 5-year EFS (pEFS) for the whole group of patients increased from 36 to 47%. In standard- and high-risk groups, the 5-year pEFS was 62 and 33%, respectively. The probability of 5-year disease-free survival was 58% in the whole group, and there were no differences between risk groups. Unsatisfactory treatment results in children classified into the high-risk group are principally due to the low remission rate.

In vitro sensitivity of leukemic cells to nucleoside derivatives in childhood acute leukemias: good activity in leukemic relapses.

Nucleoside analogues such as fludarabine and cladribine are used in therapy of indolent lymphomas and leukemias in adults, while cytarabine is used mainly in protocols for acute leukemias. Mechanisms of their activity is based on inhibition of enzymes involved in DNA, RNA and protein synthesis. The objective of the study was the analysis of in vitro cellular drug sensitivity in childhood acute lymphoblastic (ALL) and myeloid (AML) leukemia. Isolated leukemic cells obtained from 264 patients, including 152 initial ALL, 45 relapsed ALL, 54 initial AML and 13 relapsed AML were tested for cytotoxicity for fludarabine, cladribine, and cytarabine by the MTT assay. Drug concentration lethal to 50% of tested cells was regarded as a value of drug resistance. Three tested nucleoside analogues showed highest cytotoxicity against initial ALL samples. Samples of relapsed ALL and initial AML were more resistant than ALL de novo ones. Unexpectedly, no differences were observed between initial and relapsed AML samples for all tested drugs, what suggests that nucleoside analogues are active drugs in relapsed AML, which is commonly regarded as a resistant disease. All tested drugs presented significant cross-resistance in each of analyzed subgroups. In summary, tested nucleoside analogues presented relatively good activity against childhood leukemias at relapse stage.

Skeletal status in survivors of acute lymphoblastic leukemia assessed by quantitative ultrasound at the hand phalanges: a longitudinal study.

The skeletal growth in a course of acute lymphoblastic leukemia (ALL) may be affected, and the aim of the longitudinal study was to assess the skeletal status in survivors of (ALL). The studied population consisted of 38 subjects (17 female and 21 male) measured at the age of 13.9 +/- 3.8 years (5.7 +/- 2.9 years after completion of the therapy, 11.0 +/- 14.4 years after diagnosis) and 2 years earlier; compared with 1402 controls (628 female and 774 male). Patients and controls did not differ significantly in regard to age, height or weight. Skeletal status was assessed by quantitative ultrasound (US) measurements at the hand phalanges using the DBM Sonic 1200, which measures amplitude-dependent speed-of-sound, Ad-SoS (m/s). rms CV% was 0.43%. Mean baseline Ad-SoS value in patients was 1990 +/- 76 m/s and, at second measurement, 2045 +/- 86 m/s (p < 0.000001). In 31 patients, Ad-SoS increased and, in one patient, decreased more than the value of the least significant change. In controls, mean Ad-SoS values were 1973 +/- 64 m/s (baseline) and 2016 +/- 86 m/s (follow-up) and did not differ significantly vs. baseline values in patients. At second measurement, Ad-SoS in controls was significantly lower than in patients (p < 0.05). In five patients with low baseline Ad-SoS values, bone mineral density (BMD) at the spine using DPX-L was estimated; baseline mean BMD was 0.95 +/- 0.11 g/cm2, Z-score was 1.25 +/- 0.97 and, at second measurement, 1.16 +/- 0.07 g/cm2, Z-score was 0.23 +/- 0.43. A significant increase in BMD (p < 0.01) and Z-score (p < 0.05) was noted. In patients, Ad-SoS correlated significantly with age, period after completion of the therapy, body size and Tanner stages (r ranged from 0.43 to 0.83, p ranged from 0.0001 to 0.05). It can be concluded that skeletal status assessed by quantitative US at the hand phalanges in survivors of ALL improved significantly over the period of observation.

Skeletal status in survivors of childhood acute lymphoblastic leukemia assessed by quantitative ultrasound: a pilot cross-sectional study.

Intensive treatment in acute lymphoblastic leukemia (ALL) with the use of multimodality therapies, including radiotherapy, corticosteroids and cytotoxic agents, may lead to disturbances in bone metabolism. The aim of this study was to determine the degree of possible changes in bone status in survivors of ALL. The studied population consisted of 54 subjects aged 13.0 +/- 3.3 years (31 girls and 23 boys). The mean age at diagnosis was 5.5 +/- 3.5 years, age at the completion of therapy was 8.4 +/- 3.5 years, and the period of follow-up was 4.6 +/- 3.4 years. Patients were divided into three subgroups (low-, moderate- and high-risk) according to the presence of risk factors of ALL and compared with 1020 healthy subjects (508 girls and 512 boys). Patients and controls did not differ significantly in regard to age, height, or weight. Bone status was assessed by quantitative ultrasound (US) at right (dominant) hand phalanges using DBM Sonic 1200 (IGEA, Carpi, Italy) that measures amplitude-dependent speed of sound (Ad-SoS, m/s). Root mean square (RMS)_CV% was 0.43%. Mean Ad-SoS values were 2018 +/- 73 m/s in patients and 2003 +/- 80 m/s in controls and did not differ significantly. Ad-SoS tended to be lower in moderate- and high-risk patients, but differences were not significant. Ad-SoS correlated significantly with age in patients (r value ranged from 0.63 to 0.77, p < 0.01) and controls (r value ranged from 0.79 to 0.84, p < 0.0001). In multiple forward regression analysis, the following equation was obtained: Ad-SoS (m/s) = 1878 (m/s) + 11.4 x age at the study (y) + 4.0 x period after therapy completion (y) - 9.5 x duration of the therapy (y). It can be concluded that bone status assessed by quantitative US at the hand phalanges in survivors of acute lymphoblastic leukemia 4.6 years after completion of the therapy is not affected in comparison to healthy controls.

In vitro comparative antileukemic activity of various glucocorticoids in childhood acute leukemia.

Resistance to glucocorticoids is nowadays one of the strongest adverse risk factors in the treatment of childhood acute lymphoblastic leukemia (ALL). Differential in vitro antileukemic activity of various glucocorticoids and their cross-resistance pattern in childhood acute lymphoblastic and myeloblastic leukemia was determined by means of the MTT assay in 49 successfully tested samples of childhood acute leukemia. The equivalent antileukemic concentrations of respective drugs against lymphoblasts in de novo ALL samples were: 35 microM of hydrocortisone; 8 microM of prednisolone; 1.6 microM of methylprednisolone; 0.47 microM of dexamethasone and 0.23 microM of betamethasone. In comparison to initial ALL samples, the group of relapsed ALL was more resistant to: prednisolone (38-fold, p=0.004), dexamethasone (>32-fold, p=0.004), methylprednisolone (37-fold, p=0.039), betamethasone (38-fold, p=0.018) and hydrocortisone (33-fold, p=0.030). The group of acute myeloid leukemia (AML) samples were resistant to: prednisolone (>83-fold, p<0.001), dexamethasone (>32-fold, p<0.004), methylprednisolone (>65-fold, p=0.003), betamethasone (>66-fold, p=0.004) and hydrocortisone (61-fold, p=0.007), when compared to ALL at presentation. A significant cross-resistance between all used glucocorticoids as well as between glucocorticoids and other tested anti-leukemic drugs was found. In some individual cases in vitro glucocorticoid cross-resistance was less pronounced and relatively good antileukemic activity of betamethasone was observed.

Expression of CD20 on acute lymphoblastic leukemia cells in children.

CD20 determinant expressed on B precursors is associated with regulation of proliferation, apoptosis and maturation of these cells. The acute lymphoblastic leukemia "common" type (cALL) based on expression of CD20 is subdivided in type I and II. However, the clinical significance of CD20 expression on cALL and significance of cALL type I and II discernment are not fully elucidated. The association of CD20 expression with the expression of multidrug resistance molecule (MDR), CD34, atypical immunophenotypes of leukemia cells and response to induction therapy were determined in the group of 147 patients with acute lymphoblastic leukemia (ALL) B progenitor type (ALL-proB -14 patients) and common type (cALL-133 patients). The expression of CD20 on leukemia cells was studied routinely at diagnosis before the therapy. This expression was noted on leukemia cells of 6 ALL-proB patients (42.8%) and 66 cALL patients (49.6%). The expression of CD20 showed no association with the expression of CD34, CD22 and MDR. The reverse association was observed between CD20 expression and the presence of co-expression of myeloid (CD13, CD33, CD65, CD15) and T lymphoid determinants (CD2, CD5, CD7) on leukemia cells. The effect of induction therapy analyzed as time of blast cells cytoreduction in peripheral blood and time of reaching the complete remission showed the slower clearance of peripheral blood from blast cells associated with expression of CD20. There was no association of CD20 expression with the time of reaching the hematological remission. The above results suggested a "protective" role of CD20 against co-expression of other determinants (myeloid and lymphoid) and no association with the results of induction therapy.

[Recurrent pneumothorax in a child with Langerhans' cell histiocytosis]. / Nawracajaca odma oplucnowa u dziecka chorego na histiocytoze z komórek Langerhansa.

We are presenting a 16 months old boy, who has been previously developing normally, physically very active, and who suddenly developed right-sided pneumothorax with infiltrations in both lungs, initially defined as inflammatory. After placing suction drainage of the pleural cavity and antibiotic administration the child's condition improved quickly. The infiltration changes still remained in lungs arousing a suspicion of fibrocystic changes. During an attempt at changing the drainage into a water one, the symptoms of pneumothorax with hypertension increased again. A minithoracotomy with an edge resection of segment 4 and pleurectomy were performed, relating to the histopathological test of the sample, Largenhans' cell histiocytosis was diagnosed. The symptoms of diabetes insipidus appeared. The treatment according to the program DAL-HX83/90 modified by the Polish Group for Leukaemia and Malignant Lymphoma was introduced. During the treatment inducing remission a pneumothorax occurred two more times.

[Deafness, as a complication of Pseudomonas aeruginosa septicemia in a 9-year old boy with acute lymphoblastic leukemia]. / Gluchota, jako powiklanie posocznicy wywolanej paleczka ropy blekitnej u 9-letniego chlopca chorujacego na ostra bialaczka limfoblastyczna.

We described natural history of Pseudomonas aeruginosa septicemia in 9 year old boy, who was treated for acute lymphoblastic leukemia (ALL). After 14 day treatment of ALL the following signs and symptoms occurred: fever, earache with otorhoea, deafness, bilateral peripheral paralysis of n. VII, erythema, pneumonia, paralytic ileus. After 4 weeks of antimicrobial and supportive therapy, in the 10th week of chemotherapy, he achieved haematological remission. During continuation therapy, two-stage bilateral myringoplasty was performed. At present the maintenance therapy is continued, and in the future hearing aid and cochlear implant, will be applied.

[Mesenteric lipoblastoma in 13-month-old boy]. / Lipoblastoma krezki u 13-miesiecznego chlopca.

A case of lipoblastoma of mesentery in 13-month-old normally growing up boy was presented. During routine examination due to upper respiratory airways viral infection large abdominal tumor was found. Preoperative diagnostics let us establish its the most probable macro- and microscopic character and the tissue limits. Radical operation was performed. During the 5-month follow up period after the surgery there were not evident complications, we did not find any signs of tumor recurrence.

[Exercise tolerance in patients after acute lymphoblastic leukemia treatment in childhood]. / Wydolnosc wysilkowa pacjentów po leczeniu ostrej bialaczki limfoblastycznej w wieku dzieciecym.

The purpose of the presented study was to define the exercise tolerance in patients after acute lymphoblastic leukemia (ALL) treatment in childhood. Three groups of persons were examined: group A: 20 children, aged 7-19 years (mean 12.4 y), examined immediately after ALL therapy completion, with cumulative anthracycline (ATC) doses administered 155.8-300 mg/m2 and dexrazoxane, as cardioprotectant, group B: 36 patients, aged 12-24 years (mean 15.9), being 3-5 years after ALL treatment, who received ATC in cumulative doses 148.6-416.7 mg/m2, without cardioprotection, group C: 28 healthy volunteers, aged 9-25 years (mean 17.3), as controls. All the examined patients belonged to NYHA functional class I. In all subjects the exercise treadmill test was performed according to modified Bruce protocol. The parameters analysed were: MET--number of metabolic effort units achieved at the test, HRmax--maximal heart rate during exercise, %HRmax--percent of maximal HR for given patient's age achieved during the STdep--depression of ST segment in electrocardiography (ECG) immediately after the maximal exercise. During the exercise members of all 3 groups achieved the required HRmax without serious complaints and ECG abnormalities. Examined persons in group A,B and C presented with effort levels (MET), %HRmax, STdep that did not differ significantly. Only HRmax in groups A and C were higher than that achieved by members of group B.

[Lateral cervical cyst in 13-year-old girl]. / Torbiel boczna szyi u 13-letniej dziewczynki.

We present a clinical description of a 13-year-old girl with lateral cervical cyst. We particularly emphasize the need for ultrasonographic examination during routine diagnostics of so-called cervical tumors.

[Extramedullary (bone) relapse 16 months after allo bmt in a 7-year-old girl treated for acute lymphoblastic leukaemia from the second year of life]. / Pozaszpikowa (kostna) wznowa u 7-letniej dziewczynki leczonej od drugiego roku zycia z powodu ostrej bialaczki limfoblastycznej--W 16. Miesiacu po allo bmt.

7-year-old girl treated for acute lymphoblastic leukaemia since June 1995 after marrow relapse (June 1997) presented with painful oedema of the upper part of left hip and limping, 16 months after allogeneic bone marrow transplantation (BMT). Bone marrow examination excluded medullary relapse. Histopathological investigation of periosteum and bone scrapings revealed massive leukemic infiltration. Radiotherapy resulted in local arrest of the malignant process. However, bone marrow relapse was diagnosed in the child two months later with subsequent death after one month.

[Treatment regimen for children and adolescents with Hodgkin's disease designed to decrease late complications of radiotherapy]. / Program leczenia choroby hodgkina u dzieci i mlodziezy zmierzajacy do ograniczenia powiklan po radioterapii.

Between 1997 to 1999 in 9 centres of the Polish Paediatlic Leukemia/Lymphoma Study Group, 167 children and adolescents (aged 2-19 years) with stage 1 to IV Hodgkin's disease (HD) were treated according to a regimen with a limited use of radiotherapy (RT). All patients received B-DOPA and MVPP chemotherapy. The number of cycles of chemotherapy was adjusted in respective risk groups. In 13 children with stage IA and IIA disease with favourable prognostic factors chemotherapy alone was used. In other patients the dose of RT applied to lymphatic regions was 15-46,4 Gy. In case of a small tumour at presentation and good response to initial chemotherapy the RT dose was 15-16 Gy. In other cases doses of 25-30 Gy were planned. The use of higher doses, particularly exceeding 35 Gy, in eleven patients, was not justified. Among all the 167 patients, three oftliem (1.2%) with advanced disease (Stage III-1V) did not achieve first remission. The 4-year overall survival (OS), relapse free survival (RFS) and event free survival (EPS) were 99%. 93% and 90%, respectively. Relapses occurred in 8 children (first remission lasted for 4-29 (median = 9 months). All 13 children in whom chemotherapy alone was used remain in first remission. In the group of children who received RT in the dose of 15-16 Gy relapse occurred in one child. Our preliminary analysis indicates that limited use of RT in selected cases of HD in children and adolescents did not show worse results of treatment. However, the assessment of possible influence of this regimen on the decreased rate of late complications requires longer follow-up.

[High risk acute lymphoblastic leukaemia in children. Preliminary report after introducing a new version of New York (1997) protocol adjusted to the age of the patients. Report of the Polish Paediatric Leukaemia/Lymphoma Study Group]. / Ostra bialaczka limfoblastyczna u dzieci w grupie duzego ryzyka. Wstepne obserwacje po wprowadzeniu kolejnej modyfikacji (1997) protokolu Nowy York dostosowanej do wieku. Raport Polskiej Pediatrycznej Grupy ds. Leczenia Bialaczki i Chloniaków.

The paper presents the experience of the Polish Paediatric Leukaemia/Lymphoma Study Group in the treatment of high-risk acute lymphoblastic leukaemia in children using a new version of the New York (1997-1999). Protocol with treatment intensity adjusted according to the age of the patients. From April 1997 to December 1999 a group of 49 children with leukocytosis ranging from 50 900/mm3 to 580 000/mm3 (median 122 000/mm3) and 6 children with leukocytosis below 50 000/mm3 and poor response to steroids were treated with this protocol. Children below 10 years (43 patients) were treated according to the previous protocol, children above 10 years (12 patients) were treated with intensified protocol (high doses of ARA-C in consolidation and intermediate doses of Mtx in maintenance). Induction was identical for all patients. Complete remission was achieved in 92.6% patients. There were 2 relapses. Six children died - 3 without remission, 2 due to a relapse, 1 due to treatment complications. The current opinions concerning classification of HRG-ALL and treatment possibilities in this group of children are discussed.

[Intermediate doses of cytosine arabinoside in the treatment of acute non-lymphoblastic leukaemia in children]. / Sredniowysokie dawki arabinozydu cytozyny w leczeniu dzieci z ostra bialaczka nielimfoblastyczna.

Between 1995-1997, at 7 centres of the Polish Paediatric Leukaemia/Lymphoma Study Group (PPLLSG) treatment was started in 102 children with acute non-lymphoblastic leukaemia. Sixty-two children treated according to the new protocol adjusted for risk factors were evaluated. Thirty-one patients belonged to standard risk and 23 to high risk group. Eight children were not evaluated due to early death. Out of 62 children, 44 (70,9%) achieved remission; in standard and high risk groups the rates of remission were 87,5% and 73%, respectively. Four-year event-free survival (EFS), relapse-free survival (RFS) and overall survival (OS) probability in all patients were: 40,2%, 42% and 59% respectively, in standard risk group: 49,5%, 52,5%, 59,1%; in high risk group: 42%, 43,4% and 57,8%. In comparison with the previous period (1983-1994) EFS increased from 30% to 42%, which was statistically insignificant.

[Results of treatment of children with chronic myelogenous leukaemia (CML) obtained by the Polish Paediatric Leukaemia/Lymphoma Study Group]. / Wyniki leczenia dzieci chorych na przewlekla bialaczke, szpikowa (CML) w materiale Polskiej Pediatrycznej Grupy ds Leczenia Bialaczek i Chloniaków u Dzieci.

Retrospective analysis of 102 children with CML from 9 paediatric centres in Poland has been performed. A total number of 102 children: 58 boys and 44 girls aged 1-17 years (median 9.4 years old) with CML, treated in the period 1975-1999 were included in the study. Forty eight of 102 (47.1 %) children were treated with cytostatic drugs without IFN alpha: busulfan, hydroxyurea, 6-mercaptopurine or etoposide (VP-16). Fifty four of 102 (52.9%) patients were treated with interferon alpha (IFN alpha) after cytoreductive pretreatment. Thirty out of 102 (29.4%) patients underwent stem cell transplantation (SCT): 24 - matched related donor allo-BMT, 2 - matched unrelated donor allo-BMT, 1 - partially matched related donor T-cell depleted allo-PBPCT, 1 - syngeneic allo-BMT and 2 - autologous PBPCT. Overall survival analysis revealed that 46 of the 102 (45.1%) children remained alive: 5/35 (14.3%) children treated with cytostatics alone, 22/37 (59.5%) children treated with IFN alpha and 19/30 (63.3%) children treated with SCT. Among SCT survivors there are 10/17 (58.8%) children treated with IFN alpha prior to SCT and 9/13 (69.2%) children treated with cytostatics alone prior to SCT. The probability of 5-year survival is 0.51 in the group treated with SCT (median follow-up 58 months); 0.43 in the group treated with IFN alpha (median follow-up 53 months) and 0.23 in the group treated with cytostatics (median follow-up 31 months). Our data show, that BMT is the treatment of choice in CML in children. IFN alpha could be successfully applied as an alternative treatment for those, who do not have a suitable donor for allogeneic SCT. Better outcome in post BMT children, who were not treated with IFN alpha prior to SCT requires confirmation by studies on larger groups of patients. However, it seems to be justified to stop IFN alpha therapy at least 3 months before SCT. The main reason for unsuccessful treatment outcome in patients with CML in Poland remains the still insufficient access to MUD-BMT.

[Preliminary results of BFM 96 protocol in treatment of childhood ALL relapse in the experience of the Polish Paediatric Leukaemia /Lymphoma Study Group]. / Wstepne wyniki leczenia i nawrotu ostrej bialaczki limfoblastycznej cobl wg programu BFM 96 u Dzieci w Materiale Polskiej Grupy ds. Leczenia Bialaczek i Chloniaków.

Between 1998 and 1999, 36 children aged from 3 months to 18 years (10 girls and 26 boys) with first relapse of acute lymphoblastic leukaemia were included in the study. The children were treated according to the BFM 96 relapse protocol. There were 24 cases with early (including 9 children with very early) and 12 cases with late relapse ( BM-20, local extra BM-6, combined 10). The overall second complete remission (CR) rate was 83,33%. The probability of overall EFS after 2 years was 73,3%. The results obtained with BFM 96 chemotherapy in children with first late relapse are acceptable. For children with early relapses, megachemotherapy with BMT in second remission should be used.

[Adverse reactions associated with the use of L-asparaginase during therapy of patients with nB non-Hodgkin's lymphoma. Report of the Polish Paediatric LeuKaemia/Lymphoma Study Group]. / Objawy niepozadane leczenia preparatami L-asparaginazy u chorych z nieziarniczym chloniakiem zlosliwym typu NB w oparciu o material Polskiej Pediatrycznej Grupy ds. Leczenia Bialaczek i Chloniaków Zlosliwych u dzieci.

The aim of the study was to determine the side effects of asparaginase administration during treatment protocol for childhood non-Hodgkin's lymphoma (NHL). Drug adverse reactions occurred in 20/66 of patients (30,3%) treated in 9 centres in Poland between 1993 and 1998. The most common side effects were coagulation disturbances in 12/66 of the children (18,2%), which occurred due to reduced production of important coagulation factors. Six patients (9,1%) developed impairment of liver function (9,1%). Drug toxicity caused the modifications of treatment protocol in 12/66 (18,2%) of patients, mainly in the induction phase; 3 children died due to relapse of disease.