Matthew J. Barth

Genomewide linkage analysis of quantitative spirometric phenotypes in severe early-onset chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is a common, complex disease associated with substantial morbidity and mortality. COPD is defined by irreversible airflow obstruction; airflow obstruction is typically determined by reductions in quantitative spirometric indices, including forced expiratory volume at 1 s (FEV(1)) and the ratio of FEV(1) to forced vital capacity (FVC). To identify genetic determinants of quantitative spirometric phenotypes, an autosomal 10-cM genomewide scan of short tandem repeat (STR) polymorphic markers was performed in 72 pedigrees (585 individuals) ascertained through probands with severe early-onset COPD. Multipoint variance-component linkage analysis (using SOLAR) was performed for quantitative phenotypes, including FEV(1), FVC, and FEV(1)/FVC. In the initial genomewide scan, significant evidence for linkage to FEV(1)/FVC was demonstrated on chromosome 2q (LOD score 4.12 at 222 cM). Suggestive evidence was found for linkage to FEV(1)/FVC on chromosomes 1 (LOD score 1.92 at 120 cM) and 17 (LOD score 2.03 at 67 cM) and to FVC on chromosome 1 (LOD score 2.05 at 13 cM). The highest LOD score for FEV(1) in the initial genomewide scan was 1.53, on chromosome 12, at 36 cM. After inclusion of 12 additional STR markers on chromosome 12p, which had been previously genotyped in this population, suggestive evidence for linkage of FEV(1) (LOD score 2.43 at 37 cM) to this region was demonstrated. These observations provide both significant evidence for an early-onset COPD-susceptibility locus on chromosome 2 and suggestive evidence for linkage of spirometry-related phenotypes to several other genomic regions. The significant linkage of FEV(1)/FVC to chromosome 2q could reflect one or more genes influencing the development of airflow obstruction or dysanapsis.

Rituximab and FAB/LMB 96 chemotherapy in children with Stage III/IV B-cell non-Hodgkin lymphoma: a Children’s Oncology Group report

Rituximab and FAB/LMB 96 chemotherapy in children with Stage III/IV B-cell non-Hodgkin lymphoma: a Children’s Oncology Group report

Ofatumumab demonstrates activity against rituximab-sensitive and -resistant cell lines, lymphoma xenografts and primary tumour cells from patients with B-cell lymphoma

Ofatumumab is a new monoclonal antibody (mAb) targeting a novel membrane‐proximal epitope on CD20. To better define ofatumumab’s activity, we conducted pre‐clinical studies in rituximab‐sensitive cell lines (RSCL), rituximab‐resistant cell lines (RRCL), ofatumumab‐exposed cell lines (OECLs), primary lymphoma cells, and a lymphoma xenograft model. RRCL and OECL were generated by repeated exposure of sensitive cells to escalating doses of rituximab or ofatumumab ± human serum. Antibody‐dependent cellular cytotoxicity (ADCC) and complement‐mediated cytotoxicity (CMC) assays were performed to assess cellular sensitivity to rituximab or ofatumumab. Ofatumumab elicited a higher rate of CMC in RSCL, RRCL and primary tumour cells. The chronic exposure of lymphoma cells to ofatumumab resulted in rituximab resistance but less ofatumumab resistance. In an in vivo severe combined immunodeficiency mouse model of human lymphoma, ofatumumab prolonged median survival compared to rituximab. While rituximab CMC diminished with CD20 down‐regulation in RRCL passages, ofatumumab activity in vitro diminished to a lesser degree. Our data suggest that ofatumumab is more potent than rituximab in rituximab‐sensitive or rituximab‐resistant models and has the potential to decrease the development of biological resistance in patients with repeated exposure to anti‐CD20 mAbs.

Rituximab with chemotherapy in children and adolescents with central nervous system and/or bone marrow-positive Burkitt lymphoma/leukaemia: a Children's Oncology Group Report

Children and adolescents with Burkitt Lymphoma (BL) and combined central nervous system (CNS) and bone marrow involvement still have a poor prognosis with chemotherapy alone. We therefore investigated in children and adolescents with bone marrow (≥25% blasts) and/or CNS‐positive Burkitt lymphoma the chemoimmunotherapy combination of rituximab (375 mg/m2) and the standard chemotherapy arm of our previously reported French‐American‐British (FAB) Lymphome Malins de Burkitt (LMB) 96 trial. Central pathological and cytogenetic characterization was also performed. There were 40 evaluable patients with Burkitt histology (25 with leukaemia and 15 with CNS disease ± leukaemia). The chemoimmunotherapy regimen was well tolerated. The incidence of grade III/IV mucositis during induction cycles with combined chemotherapy and rituximab was 31% and 26%, respectively. The 3‐year event‐free survival (EFS)/overall survival (OS) was 90% (95% confidence interval [CI], 76–96%) in the entire cohort and 93% (95% CI, 61–99%) in patients with CNS disease. Based on the results of this trial, an international randomized study of FAB/LMB 96 chemotherapy ± rituximab for high‐risk patients is currently under investigation.

Targeting CD20+ Aggressive B-cell Non–Hodgkin Lymphoma by Anti-CD20 CAR mRNA-Modified Expanded Natural Killer Cells In Vitro and in NSG Mice

Chu and colleagues show that K562-mbIL15-41BBL–expanded peripheral blood NK cells, modified with mRNA nucleofection of an anti-CD20 CAR, significantly enhanced cytotoxicity against CD20+B-cell non-Hodgkin lymphoma, extended survival time, and reduced tumor size in xenografted NSG mice. The prognosis is very dismal for patients with relapsed CD20+ B-cell non-Hodgkin lymphoma (B-NHL). Facilitating the development of alternative novel therapeutic strategies is required to improve outcomes in patients with recurrent/refractory CD20+ B-NHL. In this study, we investigated functional activities of anti-CD20 CAR-modified, expanded peripheral blood NK cells (exPBNK) following mRNA nucleofection against CD20+ B-NHL in vitro and in vivo. CAR+ exPBNK had significantly enhanced in vitro cytotoxicity, compared with CAR− exPBNK against CD20+ Ramos (P < 0.05), Daudi, Raji, and two rituximab-resistant cell lines, Raji-2R and Raji-4RH (P < 0.001). As expected, there was no significant difference against CD20− RS4;11 and Jurkat cells. CD107a degranulation and intracellular IFNγ production were also enhanced in CAR+ exPBNK in response to CD20+ B-NHL–specific stimulation. In Raji-Luc and Raji-2R-Luc xenografted NOD/SCID/γ-chain−/− (NSG) mice, the luciferase signals measured in the CAR+ exPBNK-treated group were significantly reduced, compared with the signals measured in the untreated mice and in mice treated with the CAR− exPBNK. Furthermore, the CAR exPBNK-treated mice had significantly extended survival time (P < 0.001) and reduced tumor size, compared with those of the untreated and the CAR− exPBNK-treated mice (P < 0.05). These preclinical data suggest that ex vivo–exPBNK modified with anti-CD20 CAR may have therapeutic potential for treating patients with poor-risk CD20+ hematologic malignancies. Cancer Immunol Res; 3(4); 333–44. ©2014 AACR.

Obinutuzumab (GA101) compared to rituximab significantly enhances cell death and antibody-dependent cytotoxicity and improves overall survival against CD20(+) rituximab-sensitive/-resistant Burkitt lymphoma (BL) and precursor B-acute lymphoblastic leukaemia (pre-B-ALL): potential targeted therapy in patients with poor risk CD20(+) BL and pre-B-ALL.

Obinutuzumab is a novel glycoengineered Type‐II CD20 monoclonal antibody. CD20 is expressed in approximately 100% of children and adolescents with Burkitt lymphoma (BL) and 40% with precursor B‐cell acute lymphoblastic leukaemia (pre‐B‐ALL). We evaluated the anti‐tumour activity of obinutuzumab versus rituximab against rituximab‐resistant (Raji 4RH) and ‐sensitive (Raji) BL and pre‐B‐ALL (U698‐M) cells in vitro and in human BL or Pre‐B‐ALL xenografted mice. We demonstrated that obinutuzumab compared to rituximab significantly enhanced cell death against Raji 35·6 ± 3·1% vs. 25·1 ± 2·0%, (P = 0·001), Raji4RH 19·7 ± 2·2% vs. 7·9 ± 1·5% (P = 0·001) and U‐698‐M 47·3 ± 4·9% vs. 23·2 ± 0·5% (P = 0·001), respectively. Obinutuzumab versus rituximab also induced a significant increase in antibody‐dependent cellular cytotoxicity (ADCC) with K562‐IL15‐41BBL expanded NK cells against Raji 73·8 ± 8·1% vs. 56·81 ± 4·6% (P = 0·001), Raji‐4RH 40·0 ± 1·6% vs. 0·5 ± 1·1% (P = 0·001) and U‐698‐M 70·0 ± 1·6% vs. 45·5 ± 0·1% (P = 0·001), respectively. Overall survival in tumour xenografted mice receiving 30 mg/kg of obinutuzumab was significantly increased when compared to those receiving 30 mg/kg of rituximab in BL; Raji (P = 0·05), Raji4RH (P = 0·02) and U698‐M (P = 0·03), respectively. These preclinical data suggest obinutuzumab is significantly superior to rituximab in inducing cell death, ADCC and against rituximab‐sensitive/‐resistant BL and pre‐B‐ALL xenografted mice. Taken together, these preclinical results provide evidence to suggest that future investigation of obinutuzumab is warranted in patients with relapsed/refractory CD20+ BL and/or pre‐B‐ALL.

The future role of monoclonal antibody therapy in childhood acute leukaemias

Leukaemia is the single most common childhood malignancy. With modern treatment regimens, survival in acute lymphoblastic leukaemia (ALL) approaches 90%. Only about 70% of children with acute myeloid leukaemia (AML) achieve long term survival. Patients who relapse have a dismal prognosis. Novel therapeutic approaches are needed to improve treatment outcomes in newly‐diagnosed patients with a poor prognosis and for patients with relapsed/refractory disease that have limited treatment options. One promising approach in treating haematological malignancies has been the use of monoclonal antibodies to target cell surface antigens expressed on malignant cells. Most success with monoclonal antibody therapy in the treatment of haematological malignancies has come in the setting of adult B‐cell non‐Hodgkin lymphoma with the addition of the anti‐CD20 monoclonal antibody rituximab to standard treatment regimens. In order to further advance treatment of haematological malignancies, novel monoclonal antibodies continue to be developed that target a variety of cell surface antigens. Several antibodies continue to be investigated in childhood leukaemias. This review will discuss the development of monoclonal antibodies that target a variety of cell surface antigens for the treatment of childhood ALL and the use of the anti‐CD33 antibody gemtuzumab ozogamicin in the treatment of childhood AML.

Rituximab pharmacokinetics in children and adolescents with de novo intermediate and advanced mature B-cell lymphoma/leukaemia: a Children's Oncology Group report.

The ANHL01P1 trial was undertaken to determine pharmacokinetics and safety following the addition of rituximab to French‐American‐British/Lymphome Malins de Burkitt (FAB/LMB96) chemotherapy in 41 children and adolescents with Stage III/IV mature B‐cell lymphoma/leukaemia. Patients received rituximab (375 mg/m2) days −2 and 0 of two induction cycles and day 0 of two consolidation cycles. Highest peak levels were achieved following the second dose of each induction cycle [299 ± 19 and 384 ± 25 μg/ml (Group‐B); 245 ± 31 and 321 ± 32 μg/ml (Group‐C)] with sustained troughs and t½ of 26–29 d. Rituximab can be safely added to FAB chemotherapy with high early rituximab peak/trough levels and a long t½.

Ofatumumab: a novel, fully human anti-CD20 monoclonal antibody for the treatment of chronic lymphocytic leukemia

Ofatumumab is a fully human, IgG anti-CD20 monoclonal antibody codeveloped by GlaxoSmithKline (Brentford, UK) and Genmab (Copenhagen, Denmark). In preclinical studies, ofatumumab exhibited more potent in vitro activity than rituximab against B-cell malignancies and prolonged survival in in vivo animal models compared with rituximab. Ofatumumab is clinically well tolerated with initial infusion reactions being the predominant associated toxicity. Ofatumumab has demonstrated efficacy in relapsed/refractory chronic lymphocytic leukemia (CLL) and has received regulatory approval in both Europe and the USA for treatment of fludarabine and alemtuzumab refractory disease. Single-agent ofatumumab has resulted in overall response rates of 42-51% in relapsed/refractory CLL and up to 80% when combined with chemotherapy. In de novo CLL, overall response rates of 77-78% have been achieved.

Entinostat, a novel histone deacetylase inhibitor is active in B‐cell lymphoma and enhances the anti‐tumour activity of rituximab and chemotherapy agents

Histone deacetylases (HDACs) inhibitors are active in T‐cell lymphoma and are undergoing pre‐clinical and clinical testing in other neoplasms. Entinostat is an orally bioavailable class I HDAC inhibitor with a long half‐life, which is under evaluation in haematological and solid tumour malignancies. To define the activity and biological effects of entinostat in B‐cell lymphoma we studied its anti‐tumour activity in several rituximab‐sensitive or ‐resistant pre‐clinical models. We demonstrated that entinostat is active in rituximab‐sensitive cell lines (RSCL), rituximab‐resistant cell lines (RRCL) and primary tumour cells isolated from lymphoma patients (n = 36). Entinostat exposure decreased Bcl‐XL (BCL2L1) levels and induced apoptosis in cells. In RSCL and RRCL, entinostat induced p21 (CDKN1A) expression leading to G1 cell cycle arrest and exhibited additive effects when combined with bortezomib or cytarabine. Caspase inhibition diminished entinostat activity in some primary tumour cells suggesting that entinostat has dual mechanisms‐of‐action. In addition, entinostat increased the expression of CD20 and adhesion molecules. Perhaps related to these effects, we observed a synergistic activity between entinostat and rituximab in a lymphoma‐bearing severe combined immunodeficiency (SCID) mouse model. Our data suggests that entinostat is an active HDAC inhibitor that potentiates rituximab activity in vivo and supports its further clinical development in B‐cell lymphoma.