Intensive Polychemotherapy Research Articles

Infections Deaths in Acute Lymphoblastic Leukemia More Frequent in High Risk Leukemia

Abstract Background Acute lymphoblastic leukemia (ALL) in children currently has more than 90% survival in higher resource countries. However, in less resource countries, the survival rates are lower and infections are frequent causes. Risks for infections and poor outcomes vary during the chemotherapy phases. There is a paucity of data concerning the frequencies of the chemotherapy phase where the event is more probable. Our primary objective is to describe infections frequencies and outcomes of infection in children and also to look for association with risk factors. Methods Patients (Pts) were treated according to ALLIC-BFM-2010 protocol, a simpler branch of the BFM-AEIOP protocol and accrued prospectively. The main question is comparing outcomes of two randomizations using higher dose chemotherapy based on the leukemia risk. The risk severity was based on levels of measurable residual disease (<0.1, 0.1-10, >10 %,) and categorized in high (HR), intermediate (IR), and low (LR) risks. In SR Pts with B-cell-precursor ALL two doses of daunorubicin were prescribed in induction compared with four doses in all others. In consolidation, high-dose methotrexate was administered at 5g/m2 for SR/IR T-cell ALL and at 2g/m2 for SR/IR B-precursor-ALL. Consolidation for HR pts consisted of three intensive polychemotherapy blocks. Only T-ALL and HR pts age > 1 year received prophylactic cranial radiotherapy (12 Gy). Therapeutic cranial radiotherapy was reserved for pts with initial CNS. We performed a descriptive analysis, comparison of frequencies and proportions using Fisher test/Ji2. For time to event proportion we used Cox Analysis. Results 2232 de novo ALL pts were analyzed. Median age 5y (IQR 3-10), 56.3% were males. 2.1% had Down Syndrome; 24% categorized as HR, 56% as IR, and 20% as LR. The Immunophenotype analysis mostly B lineage (88.8%) and less T lineage (11.2%). The majority (96.7%) of pts achieved complete remission. Estimated mean time of survival was 8.8 years (ES 0.099, CI 95% 8.38-8.77) and the overall survival rate was 74 % (0.01). Trial wide, 518 (23.2%) pts died, 59 (2.6%) pts had criteria for clinical sepsis and 459 (20%) for not related sepsis causes. According to phase of the protocol, in less than one month from diagnosis, 21 events (35.6%) were observed; from 2-3 months 9 events (15.3%), from 4-6 months 6 events, (10.2%) and afterwards 23 events (39%) (p-value 0.000). No difference was found on T vs B immunophenotype, sex, age or Down syndrome. (P-value NS). However significant differences were found on risk groups 26/538 pts (4.8%) died on HR, 24/1248 (1.9%) in IR and 9/433 (2.1%) on standard risk (p-value 0.009). There was an association between the rates of sepsis and death on pts randomized to I and II protocols (p-value 0.007 and P-value 0.007). Conclusion In this large cohort of pts, we could demonstrate association between sepsis and mortality in those high-risk groups and in those pts randomized to higher chemotherapy doses. These results will allow to establish strategies to prevent infections or maximize medical interventions preventing poor infectious outcomes in high-risk groups.

Клиническое наблюдение развития острого миелоидного лейкоза у женщины на фоне ремиссии острого лимфобластного лейкоза

The article presents a case of T-cell acute lymphoblastic leukemia (T-ALL) in a patient aged 54 years, against which acute myeloid leukemia developed. The main methods of laboratory examination are given. Intensive polychemo-therapy of T-ALL in 50% of cases causes long-term remission in adults. Treatment of acute T-cell lymphoblastic leukemia consists in long-term intensive systemic chemotherapy and intrathecal administration of chemopreparations. However, despite significant achievements in treatment of patients with this pathology, relapse of the disease occurs in 40% of patients and the risk of secondary leukemia development increases.

Clinical use of polyvalent intravenous immunoglobulins during intensive polychemotherapy in children with acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) is seen in almost 30% of cases of cancer among children. Drop in absolute neutrophil count (ANC) and immunosuppression during chemotherapy are causing the significant increase in the risk of other complications, which can lead to prolonged hospitalization, higher costs of therapy and increased mortality. The analysis concerned 78 patients treated for ALL at the Department of Pediatric Oncology, Hematology and Transplantology. The indications for the use of immunoglobulins, the regimen of administration, the dose and adverse reactions were analyzed. Intravenous immunoglobulins (IVIGs) were used in 66 (85%) of 78 patients. The standard risk group (SR) was represented by 10 (15%) patients, intermediate (IR) - 29 (44%), and high (HR) - 27 (41%). The most common were 1 and 2-day administrations - 60% and 28%, respectively, of transfusions. The spread of the IVIG doses used ranged from 43 mg to 882 mg/kg body weight. In the SR and IR groups, preparations were transfused at the reinduction stage, while in the HR-consolidation. Among the indications for IVIG, the most common was hypogammaglobulinemia-117 (42%), neutropenia-69 (25%) and infection-62 (22%). During the implementation of 279 patterns of immunoglobulin preparations, 8 (3%) post-transfusion reactions were registered. The vast majority of ALL patients required immunoglobulin substitution during polychemotherapy. The supply of preparations is safe, however, there are no unambiguous guidelines regarding their dosage.

Treatment of pediatric relapsed and refractory acute myeloid leukemia

Background. Despite the modern therapy programs including hematopoietic stem cell transplantation, the treatment outcomes for children with acute myeloid leukemia (AML) remain unsatisfactory. The 5‑year overall survival rate is about 70 %. The 5‑year overall survival rate for patients with relapsed and refractory AML is 2 times lower (about 35 %). The treatment failure rate in primary AML and unsatisfactory results in relapsed and refractory AML make it necessary to optimize therapy protocols.Aim was a long-term retro- and prospective analysis of clinical and laboratory characteristics and treatment outcomes in patients with relapsed and refractory forms of AML.Materials and methods. This article presents the treatment results of 54 patients from 1 to 18 years of age, with relapsed and refractory AML treated at the N. N . Blokhin National Medical Research Center of Oncology from 1997 to 2022.Results. A comparison of 5 different programs revealed that patients who received second remission induction with the FLA + FLA scheme had 81.8 % of response (complete or partial) achievement. Analysis of the results in achievement the second remission in patients received epigenetic agents (azacytidine, decitabine, valproic and all-trans retinoid acids) with second-line chemotherapy found that treatment response rate was 100 % (n = 27), in contrast to patients received only second-line chemotherapy (n = 27) – 81.5 % (p = 0.003). The best treatment results were in group of patients whose treatment included epigenetic agents and allogenic hematopoietic stem cell transplantation after second remission induction – 5‑year overall survival was 51.3 ± 9.7 %.Conclusion. Intensive polychemotherapy with fludarabine- and cytarabine-containing regiments with following allogeneic hematopoietic stem cell transplantation and epigenetic agents are current trend and pathogenetically based approach for relapsed and refractory pediatric AML. Probable, the definition of the role and place of targeted drugs (gemtuzumab ozogamicin) could continue the advances in treatment of such unfavorable patient group.

Embryonal rhabdomyosarcoma with distant spinal cord metastasis: case and MR-imaging

Embryonal rhabdomyosarcoma (eRMS) is one of the most common soft tissue sarcomas in children, accounting for 4.5% of all childhood tumors. Half of the eRMS occuring in the head and neck are parameningeal. About 40% of patients with eRMS can develop distant metastases. In patients with intracranial tumors, metastatic spread can occur along the central nervous system (CNS) meninges. The literature describes only 4 clinical cases of eRMS with distant metastases in the spinal cord and along the meninges. Only in two out of these four cases, CSF cytology was positive (meaning that tumor cells were detected in cerebrospinal fluid). Magnetic resonance imaging (MRI) of the central nervous system with contrast enhancement can be used to detect distant metastases in the CNS and meninges. We present a clinical case of a 4-year old girl with parameningeal eRMS. MRI of the CNS performed as part of a diagnostic check-up revealed nodal metastatic foci along the meninges of the spinal cord. In accordance with the treatment protocol, the patient was diagnosed with stage 4 disease and received intensive polychemotherapy resulting in the disappearance of the nodal lesions in the spinal cord and a good prognosis. The parents gave their consent to the use of their child's data, including photographs, for research purposes and in publications.

A clinical report of Burkitt’s lymphoma

Aim. A clinical analysis of Burkitt’s lymphoma (BL) in a 4 years-old female child.Materials and methods. A retrospective analysis was conducted for the history, disease’s course, laboratory and instrumental diagnosis and treatment in patient B. with BL, 4 years old.Results. A 4-yo patient was diagnosed with BL spread to bone marrow, CNS, lymph nodes, both kidneys and spleen. Leukocytosis in common blood profile. Elevated lactate dehydrogenase (LDH) and C-reactive protein (CRP) in biochemical blood profile. Neck lymphadenopathy, mediastinum in computed tomography (CT). Splenomegaly. Multifocal lesion of both kidneys. Retroperitoneal lymphadenopathy. Positive clinical dynamics (normalisation of body weight) is observed with background therapy, LDH 335 U/L in biochemical blood profile, reduced multifocal kidney lesion and spleen size in CT.Conclusion. A clinical case of Burkitt’s lymphoma is reported affecting the bone marrow, CNS, lymph nodes, both kidneys and spleen. Intensive polychemotherapy allowed stabilisation of the patient and containment of oncological processes.

Advanced Burkitt Lymphoma in Sub-Saharan Africa Pediatric Units: Results of the Third Prospective Multicenter Study of the Groupe Franco-Africain d'Oncologie Pédiatrique.

PURPOSETo evaluate the results of an intensive polychemotherapy regimen for Burkitt lymphoma (BL) in sub-Saharan African pediatric centers.PATIENTS AND METHODSChildren with advanced-stage BL (stages II bulky, III, and IV) treated with the GFAOP–Lymphomes Malins B (GFALMB) 2009 protocol in 7 centers between April 2009 and September 2015 were prospectively registered. Treatment regimen contained a prephase with cyclophosphamide followed by 2 induction courses (cyclophosphamide, vincristine, prednisone, high-dose methotrexate [HDMTX]), 2 consolidation courses (cytarabine, HDMTX), and a maintenance phase only for stage IV. HDMTX was given at the dose of 3 g/m2.RESULTSFour hundred patients were analyzed: 7% had stage II bulky, 76% stage III, and 17% stage IV disease. Median age was 7.3 years, and sex ratio was 1.9:1 (male:female). A total of 221 patients received the whole protocol treatment and 195 achieved complete remission (CR), 11 of them after a second-line treatment. Treatment abandonment rate was 22%. One hundred twenty-five patients died, of whom 49 deaths were related to treatment toxicity. A total of 275 patients are alive, including 25 despite treatment abandonment, but only 110 are known to be in CR with a follow-up > 1 year, indicating a high rate of loss to follow-up. Twelve-month overall survival (OS) was 60% (95% CI, 54% to 66%) and 63%, 60%, and 31%, respectively, for stage II bulky, III, and IV. Patients with stage III disease who started second induction course within 34 days had OS of 76%, versus 57% (P = .0062) beyond 34 days.CONCLUSIONThe GFA-LMB2009 protocol improved patients’ survival. Early dose intensity of treatment is a strong prognostic factor. Improving supportive care and decreasing loss to follow-up are crucial.

Leukemic phase of ALK-negative anaplastic large cell lymphoma in a patient who is on androgenic steroids: A case report.

IntroductionALK-negative anaplastic large cell lymphoma (ALCL) is a peripheral T-cell lymphoma that usually involves lymph nodes or extranodal sites. Leukemic phase of ALK-negative ALCL is exceedingly rare and often carries a poor prognosis. Androgenic steroids have gained popularity among the young, and at higher doses, it can result in immune dysregulation and may be potentially carcinogenic. Case presentation: A 30-year-old gentleman of Malay ethnicity presented to the hematology department with night fevers, loss of weight and bony pain for the past 6 weeks. He is a gymnasium instructor with a history of chronic usage of intramuscular testosterone enanthate. Physical examination revealed ecchymosis over the left elbow and hepatomegaly. A complete blood count depicted anemia, thrombocytopenia and leucocytosis. An 18-Fluorodeoxyglucose positron emission tomography (18-FDG PET/CT) imaging showed a hypermetabolic anterior mediastinal mass of 6.8 × 7.0 × 6.5 cm with diffuse hypermetabolism in the liver, spleen and axial skeleton. The bone marrow trephine and mediastinal tissue histology were consistent with leukemic ALK-negative ALCL. He was treated with CHOEP (cyclophosphamide, doxorubicin, vincristine, etoposide, prednisolone) induction chemotherapy in which he required intensive antibiotic and blood support. He progressed with worsening B symptoms and new diffuse lymphadenopathies suggesting rapid dissemination of the disease. He subsequently succumbed to multiorgan failure with disseminated intravascular coagulopathy at the intensive care unit. Conclusion: Leukemic phase ALK-negative ALCL often carries a complex karyotype and requires early intensive polychemotherapy. Use of anabolic steroids depletes the ability of defending lymphocytes to remove tumour producing cells.

Rola brentuksymabu vedotin w leczeniu chorych na opornego/nawrotowego chłoniaka Hodgkina na przykładzie dwóch opisów przypadków

Hodgkin lymphoma (HL) is a form of lymphoma with good prognosis in the majority of patients, who are cured after first line chemotherapy. However, in about 25–30% of patients, the course of the disease is unfavorable and patients require treatment of subsequent lines. In further treatment, intensive polychemotherapy is used, consolidated with hematopoietic stem cell transplantation (autologous or allogenic). Also new drugs are wanted. Immunotherapy is promising, including antibodies against CD30 — brentuximab vedotin, and antibodies, that affect the PD1/PD-L1 and PD-L2 pathways — nivolumab, pembrolizumab. Determining the optimal management of patients with relapse and refractory HL requires further investigation.

PET-guided treatment in patients with advanced-stage Hodgkin's lymphoma (HD18): final results of an open-label, international, randomised phase 3 trial by the German Hodgkin Study Group

The intensive polychemotherapy regimen eBEACOPP (bleomycin, etoposide, doxorubicin, cyclophosphamide, vincristine, procarbazine, and prednisone in escalated doses) is very active in patients with advanced-stage Hodgkin's lymphoma, albeit at the expense of severe toxicities. Individual patients might be cured with less burdensome therapy. We investigated whether metabolic response determined by PET after two cycles of standard regimen eBEACOPP (PET-2) would allow adaption of treatment intensity, increasing it for PET-2-positive patients and reducing it for PET-2-negative patients. In this open-label, randomised, parallel-group phase 3 trial, we recruited patients aged 18-60 years with newly diagnosed, advanced-stage Hodgkin's lymphoma in 301 hospitals and private practices in Germany, Switzerland, Austria, the Netherlands, and the Czech Republic. After central review of PET-2, patients were assigned (1:1) to one of two parallel treatment groups on the basis of their PET-2 result. Patients with positive PET-2 were randomised to receive six additional cycles of either standard eBEACOPP (8 × eBEACOPP in total) or eBEACOPP with rituximab (8 × R-eBEACOPP). Those with negative PET-2 were randomised between standard treatment with six additional cycles of eBEACOPP (8 × eBEACOPP) or experimental treatment with two additional cycles (4 × eBEACOPP). A protocol amendment in June, 2011, introduced a reduction of standard therapy to 6 × eBEACOPP; after this point, patients with positive PET-2 were no longer randomised and were all assigned to receive 6 × eBEACOPP and patients with negative PET-2 were randomly assigned to 6 × eBEACOPP (standard) or 4 × eBEACOPP (experimental). Randomisation was done centrally using the minimisation method including a random component, stratified according to centre, age (<45 vs ≥45 years), stage (IIB, IIIA vs IIIB, IV), international prognostic score (0-2 vs 3-7), and sex. eBEACOPP was given as previously described; rituximab was given intravenously at a dose of 375 mg/m2 (maximum total dose 700 mg). The primary objectives were to show superiority of the experimental treatment in the PET-2-positive cohort, and to show non-inferiority of the experimental treatment in the PET-2-negative cohort in terms of the primary endpoint, progression-free survival. We defined non-inferiority as an absolute difference of 6% in the 5-year progression-free survival estimates. Primary analyses in the PET-2-negative cohort were per protocol; all other analyses were by intention to treat. This trial was registered with ClinicalTrials.gov, number NCT00515554. Between May 14, 2008, and July 18, 2014, we recruited 2101 patients, of whom 137 were found ineligible before randomisation and a further 19 were found ineligible after randomisation. Among 434 randomised patients (217 per arm) with positive PET-2, 5-year progression-free survival was 89·7% (95% CI 85·4-94·0) with eBEACOPP and 88·1% (83·5-92·7) with R-eBEACOPP (log-rank p=0·46). Patients with negative PET-2 randomly assigned to either 8 × eBEACOPP or 6 × eBEACOPP (n=504) or 4 × eBEACOPP (n=501) had 5-year progression-free survival of 90·8% (95% CI 87·9-93·7) and 92·2% (89·4-95·0), respectively (difference 1·4%, 95% CI -2·7 to 5·4). 4 × eBEACOPP was associated with fewer severe infections (40 [8%] of 498 vs 75 [15%] of 502) and organ toxicities (38 [8%] of 498 vs 91 [18%] of 502) than were 8 × eBEACOPP or 6 × eBEACOPP in PET-2-negative patients. Ten treatment-related deaths occurred: four in the PET-2-positive cohort (one [<1%] in the 8 × eBEACOPP group, three [1%] in the 8 × R-eBEACOPP group) and six in the PET-2-negative group (six [1%] in the 8 × eBEACOPP or 6 × eBEACOPP group). The favourable outcome of patients treated with eBEACOPP could not be improved by adding rituximab after positive PET-2. PET-2 negativity allows reduction to only four cycles of eBEACOPP without loss of tumour control. PET-2-guided eBEACOPP provides outstanding efficacy for all patients and increases overall survival by reducing treatment-related risks for patients with negative PET-2. We recommend this PET-2-guided treatment strategy for patients with advanced-stage Hodgkin's lymphoma. Deutsche Krebshilfe, Swiss State Secretariat for Education and Research, and Roche Pharma AG.

Advances in therapy for Philadelphia-positive acute lymphoblastic leukaemia of childhood and adolescence.

The presence of the BCR/ABL1 fusion gene in childhood acute lymphoblastic leukaemia (ALL) is a rare finding and has been an adverse prognostic factor associated with a high risk of therapeutic failure. The current key components of treatment are intensive polychemotherapy and a BCR/ABL1 kinase domain inhibitor. This treatment approach has been applied in a few clinical trials by paediatric leukaemia study groups. Thus, this subtype of ALL serves as the first model system for truly targeted treatment. The role of haematopoietic stem cell transplantation (HSCT) is increasingly called into question, at least in a favourable, though not yet clearly defined, subset of patients. Currently, the choice of the most effective tyrosine kinase inhibitor is not yet settled, in particular, in view of potential reduction of overall treatment intensity.

Oral etoposide in relapsed or refractory Ewing sarcoma: a monoinstitutional experience in children and adolescents.

To assess the efficacy and toxicity of low-dose oral etoposide (VP) 16 in relapsing/refractory Ewing sarcoma. The records of all patients treated at our department between 1989 and 2012 for relapsing/refractory Ewing sarcoma who received oral VP-16 were analyzed. The dose was 40 mg/m2 daily for 21 consecutive days in every 28. Response was assessed after 2/3 cycles according to Response Evaluation Criteria in Solid Tumors 1.0. A total of 46 of 58 patients completed at least 2 cycles; 12 suspended the treatment earlier due to rapid disease progression. The patients' median age at diagnosis was 14 years and 25/58 had metastatic disease. All patients received intensive polychemotherapy including VP-16 IV as first- (n = 53) or second-line (n = 5) treatment; 21/58 had myeloablative regimens with peripheral blood stem cell rescue, and 1 underwent allogeneic stem cell transplantation. Oral VP-16 was prescribed as 2nd-, 3rd-, and 4th-line treatment for 19, 27, and 12 patients, respectively. The cycles administered totaled 241 (median 3, mean 4 per patient; range 1-14). A total of 46 of 58 patients were evaluable: 11 responded (9 partial remission, 1 very good partial remission, 1 complete remission) and 10 were stable, the response lasting a mean of 8 months. Hematologic toxicity G3/G4 (in 164/241 evaluable cycles) occurred in 15%, 16%, and 11% of cycles for leukocytes, hemoglobin, and platelets, respectively. There were 5 cases of pneumonia. Two patients developed secondary leukemia after receiving 12 and 14 cycles. Low-dose oral VP-16 may be suitable in a palliative setting with an acceptable toxicity. The risk of secondary leukemia is in line with reports in the literature.

Selenium inadequacy is not associated with oxidative stress in child and adolescent acute lymphocytic leukemia survivors

ObjectiveAcute lymphocytic leukemia (ALL) and its subsequent treatment may provoke increased oxidative stress. The aim of this study was to investigate the antioxidant status of children and adolescents who had received ALL therapy, and to test the hypothesis that selenium (Se) inadequacy is correlated with reduced defenses against oxidative stress in this population. MethodsThis case–control study involved 24 patients between ages 5 and 13 y who had been treated successfully for ALL (ALL group) and 60 children of similar age and socioeconomic background with no clinical history of leukemia (control group). Dietary intake of Se was evaluated by the 24-h recall method, and the concentrations of Se in plasma, erythrocytes, and urine determined. Antioxidant status was assessed by analysis of the oxidative stress markers, namely, superoxide dismutase (SOD), glutathione peroxidase (GPx), malondialdehyde (MDA), α-tocopherol, and 8-oxo-deoxyguanosine (8-oxo-dG). ResultsThere were no between-group differences with respect to plasma (P = 0.122), erythrocyte (P = 0.202), urinary (P = 0.608), or dietary (P = 0.757) levels of Se. GPx activity was significantly (P < 0.001) reduced in the ALL group compared with the control group, whereas SOD activity and MDA concentrations were similar. The concentrations of α-tocopherol and 8-oxo-dG were significantly increased in the ALL group compared with the control group (P < 0.001 and P = 0.031, respectively). ConclusionAll participants were Se inadequate, but such inadequacy was not correlated with reduced defenses against oxidative stress. However, individuals of the ALL group were with increased oxidative stress compared with the control group, possibly due to previous disease and to intensive polychemotherapy.

Outcomes of Innovative Programmed Treatment of Children with Malignant Loco-Motor System Tumors in Moscow

During the period from 2005 to 2010 forty nine patients from Moscow city aged 1 — 16 years (mean age 9.4 years) were treated for malignant tumors (MT) of loco-motor system. Localized process was diagnosed in 25 (51%) patients, IV stage of disease in 24 (49%) children. Use of adopted treatment program including intensive polychemotherapy, organ-saving surgical treatment and radiotherapy enabled to achieve optimistic results. Overall 2-years survival rate for children with loco-motor system MT made up 64.3±7.2%, i.e. 86.5±7.2% in localized and 42.6±10.5% in disseminated process.

Epstein-Barr Virus-Negative Aggressive Natural Killer-Cell Leukaemia with High P-Glycoprotein Activity and Phosphorylated Extracellular Signal-Regulated Protein Kinases 1 and 2

Aggressive natural killer-cell leukaemia (ANKL) is a rare type of disease with fulminant course and poor outcome. The disease is more prevalent among Asians than in other ethnic groups and shows strong association with Epstein-Barr virus (EBV) and P-glycoprotein (P-gp) expression associated with multidrug resistance. Here we present a case of a 47 year old Caucasian female with a prior medical history of azathioprine treated ulcerative colitis who developed EBV-negative form of ANKL. The patient presented with hepatosplenomegaly, fever and nausea with peripheral blood and bone marrow infiltration with up to 70% of atypical lymphoid cells positive for cCD3, CD2, CD7, CD56, CD38, CD45, TIA1 and granzyme B, and negative for sCD3, CD4, CD5, CD8, CD34 and CD123 indicative of ANKL. Neoplastic CD56+ NK-cells showed high level of P-glycoprotein expression and activity, but also strong expression of phosphorylated extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) MAP kinase. The patient was treated with an intensive polychemotherapy regimen designed for treatment of acute lymphoblastic leukaemia, but one month after admission developed sepsis, coma and died of cardiorespiratory arrest. We present additional evidence that, except for the immunophenotype, leukaemic NK-cells resemble normal NK-cells in terms of P-gp functional capacity and expression of phosphorylated ERK1/2 signalling molecule. In that sense drugs that block P-glycoprotein activity and activated signalling pathways might represent new means for targeted therapy.

Abstract 3750: The novel Akt inhibitor MK-2206, is cytotoxic in T-cell acute lymphoblastic leukemia: Therapeutic implications

Abstract T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive neoplastic disorder arising within the thymus from the clonal proliferation of T-cell precursors. T-ALL accounts for 15% of ALL cases in children and 25% in adults. Although the prognosis of T-ALL has improved, due to the use of intensive polychemotherapy schemes, the outcome of relapsed and chemoresistant T-ALL is still poor, especially in adults, with a 35-40% survival at 5 years. Therefore, efforts are being made to develop targeted therapies against deregulated signaling cascades that sustain T-ALL cell proliferation, survival, and drug-resistance. A signaling pathway that is frequently upregulated in T-ALL, is the PI3K/Akt/mTOR network. To explore whether or not Akt could represent a potential target for therapeutic intervention in T-ALL, we evaluated the effects of the novel allosteric Akt inhibitor, MK-2206, on a panel of human T-ALL cell lines (MOLT-4, CEM, drug-resistant CEM) and primary cells from T-ALL patients, characterized by pathway upregulation. MK-2206 decreased T-ALL cell line viability as documented by MTT assays. IC50 for MK-2206 ranged from 1.0 to 4.8 μM at 48 hours. The drug was effective against drug-resistant CEM cells, overexpressing 170-kDa P-glycoprotein. MK-2206 blocked leukemic cells in the G1 phase of the cell cycle and induced caspase-dependent apoptotic cell death, as documented by Annexin V/propidium iodide staining and western blot analysis. In CEM cells, MK-2206 induced autophagy, as demonstrated by an increase in the 14-kDa form of LC3A/B. Western blotting documented a concentration-dependent dephosphorylation of Akt and its downstream targets, GSK-3β and FOXO3A, in response to MK-2206. mTORC1 downstream targets were also efficiently dephosphorylated by MK-2206, including p70S6K and 4E-BP1. MK-2206 decreased mTORC2 activity, as indicated by the downregulation of Ser 2481 p-mTOR levels, a readout for mTORC2 activity In MOLT-4 and CEM cells, MK-2206 strongly synergized (combination index: 0.1-0.33) with doxorubicin. Moreover, MK-2206 dephosphorylated Akt and induced apoptosis in a T-ALL patient cell subset (CD34+/CD4−/CD7−) which is enriched in leukemia initiating cells. Our findings indicate that Akt inhibition, either alone or in combination with chemotherapeutic drugs, represents a potential therapeutic target in T-ALL cells that require upregulation of the PI3K/Akt/mTOR signaling pathway for their proliferation and survival. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3750. doi:1538-7445.AM2012-3750

Antitumor activity of new pyrazolo[3,4-d]pyrimidine SRC kinase inhibitors in Burkitt lymphoma cell lines and its enhancement by WEE1 inhibition

Burkitt lymphoma (BL) is a highly aggressive B cell neoplasm. Although intensive polychemotherapy regimens have proven very effective, they are associated with significant toxicities. Therefore, more rational therapies that selectively target the molecular abnormalities of BL are needed. Recent data suggest that the tyrosine kinase SRC could represent a therapeutic target for BL. We found that new pyrazolo[3,4-d]pyrimidine SRC inhibitors exerted a significant cytotoxic effect and induced apoptosis on two BL cell lines, as determined by MTS assays, cytofluorimetric analyses and caspase 3 assay. Notably, our SRC inhibitors proved to be more effective than the well-known SRC inhibitor PP2 [4-amino-5-(4-chlorophenyl)-7-(dimethylethyl)pyrazolo[3,4-d]pyrimidine] in BL cells. Moreover, our small molecules induced a G2/M arrest in BL cells through a possible new mechanism, whereby SRC inhibition hinders an AKT-WEE1-cyclin-dependent kinase 1 (CDK1) axis, leading to inhibition of CDK1, the main trigger of entry into mitosis. By using a small-molecule inhibitor of WEE1, a crucial CDK1 negative regulator, we were able to shift the balance toward apoptosis rather than growth arrest and enhance the efficacy of the SRC inhibitors, suggesting a possible use of these selective drugs in combination for a safe and efficient treatment of BL.

Long-Term Outcome in Children With Relapsed Acute Lymphoblastic Leukemia After Time-Point and Site-of-Relapse Stratification and Intensified Short-Course Multidrug Chemotherapy: Results of Trial ALL-REZ BFM 90

The multicenter trial ALL-REZ BFM (ie, Acute Lymphoblastic Leukemia Relapse Berlin-Frankfurt-Münster) 90 was designed to improve prognosis for children with relapsed acute lymphoblastic leukemia (ALL) by time-to-relapse- and site-of-relapse-adapted stratification and by introduction of novel chemotherapy elements and to evaluate new prognostic parameters in a large, population-based cohort. Five hundred twenty-five patients stratified into risk groups A (early bone marrow [BM] relapses), B (late BM relapses), and C (isolated extramedullary relapses) received alternating short-course intensive polychemotherapy (in blocks R1, R2, or R3) and cranial/craniospinal irradiation followed by maintenance therapy. Block R3 (high-dose cytarabine and etoposide) was introduced to improve the outcome compared with historical controls. Patients with early BM or T-ALL relapse (poor prognosis group [PPG]) were eligible for experimental regimens. One hundred seventeen patients received stem-cell transplantation (SCT). The probabilities (and standard deviations) of event-free survival (pEFS) and overall survival (pOS) at 10 years were 0.30 +/- .02 and 0.36 +/- .02, respectively. Significant differences existed between strategic groups (pEFS(A) = .17 +/- .03; pEFS(B) = .43 +/- .04; pEFS(C) = .54 +/- .06; pEFS(PPG) = .15 +/- .03; log-rank P < .001). Patients of high-risk groups A plus PPG did better with SCT than with chemotherapy (pEFS = .33 +/- .05 v 0.20 +/- .05; P = .005). The pEFS was similar to trials ALL-REZ BFM 85/87 (.36 +/- .03. v 0.37 +/- .03; P = .419; PPG excluded). Time point, site of relapse, immunophenotype, and SCT were significant predictors of pEFS in multivariate analyses. More than one third of patients in this large, population-based trial were cured. Neither R3 nor adaptation of chemotherapy intensity was capable of improving pEFS or of overcoming prognostic factors. In high-risk patients, remission induction regimens must be improved, and allogeneic SCT should be recommended in patients achieving second complete remission.

Long-term results of the Italian Association of Pediatric Hematology and Oncology (AIEOP) Studies 82, 87, 88, 91 and 95 for childhood acute lymphoblastic leukemia

We analyzed the long-term outcome of 4865 patients treated in Studies 82, 87, 88, 91 and 95 for childhood acute lymphoblastic leukemia (ALL) of the Italian Association of Pediatric Hematology and Oncology (AIEOP). Treatment was characterized by progressive intensification of systemic therapy and reduction of cranial radiotherapy. A progressive improvement of results with reduction of isolated central nervous system relapse rate was obtained. Ten-year event-free survival increased from 53% in Study 82 to 72% in Study 95, whereas survival improved from 64 to 82%. Since 1991, all patients were treated according to Berlin-Frankfurt-Muenster (BFM) ALL treatment strategy. In Study 91, reduced treatment intensity (25%) yielded inferior results, but intensification of maintenance with high-dose (HD)-L-asparaginase (randomized) allowed to compensate for this disadvantage; in high-risk patients (HR, 15%), substitution of intensive polychemotherapy blocks for conventional BFM backbone failed to improve results. A marked improvement of results was obtained in HR patients when conventional BFM therapy was intensified with three polychemotherapy blocks and double delayed intensification (Study 95). The introduction of minimal residual disease monitoring and evaluation of common randomized questions by AIEOP and BFM groups in the protocol AIEOP-BFM-ALL 2000 are expected to further ameliorate treatment of children with ALL.

Leukemic CD52 Negative Subclones Due to Defective Glycophosphatidyl-Innositol Anchoring Are Common in Acute Precursor B Lymphoblastic Leukemia, Escape Alemtuzumab Therapy, but Display Increased Sensitivity to Rituximab Mediated Complement Dependent Cytotoxicity: a Mechanistic Rationale for Antibody Combination Therapy.

Abstract 835The prognosis of B-lineage acute lymphoblastic leukemia (B-ALL) in adults is poor despite intensive polychemotherapy. Treatment intensification is limited by treatment related toxicity. Alemtuzumab (ALM) is a humanized monoclonal antibody directed against the glycophosphatidyl-innositol (GPI-) anchored membrane protein CD52. Because B-ALL frequently express CD52, ALM may be of therapeutic value in this disease. However, loss of CD52 due to acquired defects in GPI biosynthesis has been described in various hematological settings. Presence of GPI-defective leukemic subclones in ALL will have implications for the response of ALL to ALM. We analyzed leukemic cells from 24 randomly selected cases for expression of CD52 by flow cytometry using FITC-conjugated ALM. To identify GPI-defective cells counter staining was performed using PE-conjugated antibodies against the GPI-anchored surface markers CD55 and CD66c. Of the 24 cases, 19 expressed CD52 (median MFI: 220, range 34.0-617). In 16 of these 19 cases CD52− events were detected. In 11 of these 16 cases these events represented a discernable CD52−CD55−CD66c− GPI-defective subpopulation (median 0.02% of all cells, range 0.01-25.8%). We subsequently evaluated the in vivo activity of ALM against ALL, and the relevance of the GPI-defective subpopulations, in a preclinical model of human ALL. For this, we selected case BV (4.0% CD19+CD52−CD55− cells, overall MFI 137) and case CM (no detectable CD19+CD52−CD55− cells, overall MFI 616). NOD/scid mice were inoculated i.v. with primary BV or CM cells. Starting 14 days after inoculation, animals received daily i.p. injections of 250μg ALM (n=8) or saline (n=4). After 3 weeks of treatment animals were sacrificed and bone marrow (BM) was analyzed for the presence of leukemic cells by flow cytometry. BM of control treated animals engrafted with BV cells contained 80±20% leukemic cells, 95% of which were CD52+. BM of control treated animals engrafted with CM cells contained 69%±14% leukemic cells, 99% of which were CD52+. BM of ALM treated animals engrafted with BV or CM cells contained 75% (±5.9%) and 21% (±5.4%) leukemic cells, respectively. However, these cells were exclusively CD52−CD55−CD66c−, demonstrating that ALM had efficiently eradicated all CD52+cells but in both cases clonogenic GPI-defective subpopulations had been selected. The cells remained CD52−CD55−CD66c− when transplanted into secondary untreated recipients, demonstrating that the loss of expression was irreversible. As known from paroxysmal nocturnal hemoglobinuremia, GPI-defective cells are particularly susceptible to complement mediated cytotoxicity (CDC) because they lack the protective function of CD55. Because the cells recovered from ALM treated animals and control treated mice expressed similar levels of CD20, we compared their susceptibility to rituximab (RTX) mediated CDC. GPI-defective and wild type CM cells were coated with 0-10μg/mL RTX, exposed to dilutions of complement, and assayed for viability by flow cytometry. In the presence of 10μg/mL RTX, 50% lysis of wild type cells required a 1:40 complement dilution while 50% lysis of GPI-defective cells required a 1:200 dilution. Likewise, in the presence of a 1:100 complement dilution, 2.5μg/mL RTX mediated 10% lysis in wild type cells but 75% lysis in GPI-defective cells. To determine whether the GPI-defective cells that escape ALM treatment could therefore be targeted by RTX in vivo, we engrafted NOD/scid mice with CM or BV cells and administered RTX (250μg daily) or RTX plus ALM (250μg each daily) in the set-up described above. Administration of RTX alone resulted in partial reduction of engraftment levels only, most likely due to low expression levels of CD20 on the leukemic cells. However, combined administration of ALM and RTX resulted in complete disappearance of leukemic cells in 8 out of 8 CM-engrafted animals, 6 of which were in molecular remission as demonstrated by RT-PCR, and in 4 out of 5 BV-engrafted animals. We conclude that ALM is highly effective against CD52+ ALL cells but CD52− GPI-defective subclones may pre-exist in the majority of primary ALL and escape therapy. Because GPI-defective cells are particularly susceptible to complement mediated lysis, combination of ALM with antibodies directed against non-GPI anchored proteins, such as RTX, may be highly synergistic. Disclosures:No relevant conflicts of interest to declare.