Rudolf Schmits

Six versus eight cycles of bi-weekly CHOP-14 with or without rituximab in elderly patients with aggressive CD20+ B-cell lymphomas: a randomised controlled trial (RICOVER-60)

BACKGROUND Cyclophosphamide, doxorubicin, vincristine, and prednisolone (CHOP) is used to treat patients with non-Hodgkin lymphoma. Interval decrease from 3 weeks of treatment (CHOP-21) to 2 weeks (CHOP-14), and addition of rituximab to CHOP-21 (R-CHOP-21) has been shown to improve outcome in elderly patients with diffuse large B-cell lymphoma (DLBCL). This randomised trial assessed whether six or eight cycles of R-CHOP-14 can improve outcome of these patients compared with six or eight cycles of CHOP-14. METHODS 1222 elderly patients (aged 61-80 years) were randomly assigned to six or eight cycles of CHOP-14 with or without rituximab. Radiotherapy was planned to sites of initial bulky disease with or without extranodal involvement. The primary endpoint was event-free survival; secondary endpoints were response, progression during treatment, progression-free survival, overall survival, and frequency of toxic effects. Analyses were done by intention to treat. The trial is registered on National Cancer Institute website, number NCT00052936 and as EU-20243. FINDINGS 3-year event-free survival was 47.2% after six cycles of CHOP-14 (95% CI 41.2-53.3), 53.0% (47.0-59.1) after eight cycles of CHOP-14, 66.5% (60.9-72.0) after six cycles of R-CHOP-14, and 63.1% (57.4-68.8) after eight cycles of R-CHOP-14. Compared with six cycles of CHOP-14, the improvement in 3-year event-free survival was 5.8% (-2.8-14.4) for eight cycles of CHOP-14, 19.3% (11.1-27.5) for six cycles of R-CHOP-14, and 15.9% (7.6-24.2) for eight cycles of R-CHOP-14. 3-year overall survival was 67.7% (62.0-73.5) for six cycles of CHOP-14, 66.0% (60.1-71.9) for eight cycles of CHOP-14, 78.1% (73.2-83.0) for six cycles of R-CHOP-14, and 72.5% (67.1-77.9) for eight cycles of R-CHOP-14. Compared with treatment with six cycles of CHOP-14, overall survival improved by -1.7% (-10.0-6.6) after eight cycles of CHOP-14, 10.4% (2.8-18.0) after six cycles of R-CHOP-14, and 4.8% (-3.1-12.7) after eight cycles of R-CHOP-14. In a multivariate analysis that used six cycles of CHOP-14 without rituximab as the reference, and adjusting for known prognostic factors, all three intensified regimens improved 3-year event-free survival (eight cycles of CHOP-14: RR [relative risk] 0.76 [0.60-0.95], p=0.0172; six cycles of R-CHOP-14: RR 0.51 [0.40-0.65], p<0.0001; eight cycles of R-CHOP-14: RR 0.54 [0.43-0.69], p<0.0001). Progression-free survival improved after six cycles of R-CHOP-14 (RR 0.50 [0.38-0.67], p<0.0001), and eight cycles of R-CHOP-14 (RR 0.59 [0.45-0.77], p=0.0001). Overall survival improved only after six cycles of R-CHOP-14 (RR 0.63 [0.46-0.85], p=0.0031). In patients with a partial response after four cycles of chemotherapy, eight cycles were not better than six cycles. INTERPRETATION Six cycles of R-CHOP-14 significantly improved event-free, progression-free, and overall survival over six cycles of CHOP-14 treatment. Response-adapted addition of chemotherapy beyond six cycles, though widely practiced, is not justified. Of the four regimens assessed in this study, six cycles of R-CHOP-14 is the preferred treatment for elderly patients, with which other approaches should be compared.

Treatment Results in Localized Primary Gastric Lymphoma: Data of Patients Registered Within the German Multicenter Study (GIT NHL 02/96)

PURPOSE In the prospective study 02/96 on primary GI lymphoma, we have collected data on histology, clinical features, and treatment results. In particular, in stages I and II localized primary gastric lymphoma (PGL), our objectives were to reduce treatment intensity and to confirm our hypothesis from study 01/92, which maintained that an organ-preserving approach is not inferior to primary surgery. PATIENTS AND METHODS Patients receiving radiotherapy and/or chemotherapy were stratified for histologic grade, stage, and whether surgery had been carried out or not (as decided by each participating center). Patients with aggressive PGL received six cycles of CHOP-14 (cyclophosphamide, doxorubicin, vincristine, and prednisone) followed by involved-field radiotherapy (40 Gy). Patients with indolent PGL (including patients experiencing treatment failure with antibiotic therapy for Helicobacter pylori) were treated with extended-field radiotherapy. The volume depended on stage. The irradiation dose was 30 Gy, followed by a boost of 10 Gy (the latter omitted after complete resection) to the tumor region. RESULTS Seven hundred forty-seven patients were accrued. Of these patients, 393 with localized PGL were treated with radiotherapy and/or chemotherapy only or additional surgery between December 1996 and December 2003. The survival rate at 42 months for patients treated with surgery was 86% compared with 91.0% for patients without surgery. CONCLUSION In this nonrandomized study (02/96), we reproduced the previous results of study 01/92 showing no disadvantage for an organ-preserving treatment. Therefore, primary stomach resection should be questioned.

Analysis of the antibody repertoire of astrocytoma patients against antigens expressed by gliomas

The molecular characterization of antigens preferentially or exclusively expressed by astrocytomas and recognized by the autologous immune system are a prerequisite for the development of specific vaccines. To identify such antigens, we screened 5 cDNA expression libraries derived from astrocytomas and other gliomas for reactivity with high‐titered IgG antibodies in the sera of astrocytoma patients using SEREX, the serologic identification of antigens by recombinant cDNA expression cloning. Autologous and allogeneic SEREX analysis of >5 × 106 clones with the sera of 18 astrocytoma patients revealed 10 antigens: the differentiation antigen glial fibrillary acidic protein (GFAP), Bax‐inhibitor 1 (which was overexpressed in all glioma samples tested), 3 other molecules involved in the regulation of gene expression and proliferation (the nm23‐H2‐encoded nucleoside diphosphate kinase B, the Ran binding protein‐2 and a DNA binding protein encoded by the son gene), SP40,40 (a complement inhibitory molecule), the chaperonin TCP‐1, calnexin and 2 new gene products. No immune responses were detected against the “shared tumor” or “cancer testis antigens” that are regularly expressed in gliomas. Antibody responses in astrocytoma patients against antigens expressed by gliomas were rare and, with the exception of Bax‐inhibitor 1 and the product of the son gene, were also found in apparently healthy controls. We conclude that although astrocytomas express a broad spectrum of antigens, they elicit antibody responses only rarely, most likely because of their intrinsic immunosuppressive effects.

Analysis of the B cell repertoire against autoantigens in patients with giant cell arteritis and polymyalgia rheumatica

The analysis of the antibody repertoire of patients with giant cell arteritis (GCA) and polymyalgia rheumatica (PMR) might identify target antigens of the autoimmune response with potential relevance to our understanding of the pathogenesis of the disease and to the development of serodiagnostic tests. To detect such antigens, we screened a cDNA library derived from normal human testis for antigens reacting with IgG antibodies in the 1 : 250 diluted sera of three patients with untreated GCA using SEREX, the serological identification of antigens by recombinant cDNA expression cloning. Of 100 000 clones screened with each serum, six, 28 and six clones, respectively, were positive, representing a total of 33 different antigens. Most of the antigens reacted only with the serum used for identification and/or at a similar frequency with normal control sera. However, lamin C and the nuclear antigen of 14 kD reacted specifically with 32% of GCA/PMR, but with none of the control sera, while human cytokeratin 15, mitochondrial cytochrome oxidase subunit II, and a new gene product were detected preferentially, but not exclusively by sera from GCA/PMR patients. We conclude that patients with GCA/PMR develop antibodies against a broad spectrum of human autoantigens. Antibodies against human lamin C, the nuclear autoantigen of 14 kD as well as human cytokeratin 15, mitochondrial cytochrome oxidase subunit II and the product of a new gene should be investigated further to determine their value as tools for the diagnosis and/or the definition of clinical subgroups of patients with GCA/PMR.

Statins inhibit lymphocyte homing to peripheral lymph nodes.

Lymphocyte homing to peripheral lymph nodes is governed by adhesion molecules, including lymphocyte function‐associated antigen 1 (LFA‐1). Statins are 3‐hydroxy‐3‐methylglutaryl coenzyme A reductase inhibitors and exert anti‐inflammatory effects, e.g. inhibition of LFA‐1. It is still not known whether statin compounds are capable of inhibiting lymphocyte homing in vivo. We used a cervical lymph node preparation to study the effects of simvastatin on lymphocyte adhesion to high endothelial venules (HEVs) by means of intravital fluorescence microscopy (IVM). IVM revealed that firm adhesion of lymphocytes to HEV endothelium critically depends on the adhesive function of LFA‐1. The number of firmly adherent lymphocytes was reduced by 58% in LFA‐1‐deficient mice (P < 0·05 versus wild‐type controls). As in mutant mice, acute treatment with simvastatin (i.p. injection at 2 hr prior to IVM) inhibited the firm adhesion of lymphocytes to HEV endothelium of wild‐type animals by 63% (P < 0·05 versus vehicle‐treated wild‐type controls). In addition, acute treatment with the synthetic statin‐derivate LFA878 also reduced firm lymphocyte adhesion in HEVs by 63% (P > 0·05 versus placebo‐treated controls). Histological analysis after a 10‐day treatment with simvastatin showed reduced cellularity of cervical lymph nodes, as indicated by a reduction of the relative area of haematoxylin‐stained cell nuclei in cervical lymph node cross‐sections from 94 ± 0% in vehicle‐treated controls to 77 ± 3% in simvastatin‐treated mice (P < 0·05). We conclude that statin compounds are capable of inhibiting lymphocyte homing to murine peripheral lymph nodes in vivo. This may have novel implications for the treatment of adaptive immune responses, e.g. transplant rejection.

Use of Spontaneous Epstein-Barr Virus-Lymphoblastoid Cell Lines Genetically Modified to Express Tumor Antigen as Cancer Vaccines: Mutated p21 ras Oncogene in Pancreatic Carcinoma as a Model

Spontaneous Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines (SP-LCLs) can be easily obtained from latently EBV-infected cancer patients and used as a source of antigen-presenting cells (APCs) for immunotherapy. Using point-mutated (codon 12) p21(ras) (muRas) as a model tumor antigen, we evaluated the practicability of using genetically modified SP-LCLs as cancer vaccines for patients with pancreatic cancer expressing mutated Ras (muRas). The repeated stimulation of peripheral blood mononuclear cells (PBMCs) from patients with muRas-LCLs elicited a strong, muRas-specific T cell response. A significant cytotoxic activity against EBV virus proteins or components of the expression vector was not observed. The T cells were able to recognize naturally presented muRas, as shown by their cytotoxicity against muRas (Gly-12 to Val-12 or Asp-12)-expressing tumor cells. The T cell response was mainly MHC class I restricted, and peptides containing amino acids 5 to 14 of muRas-Val-12 and muRas-Asp-12 were identified as immunogenic peptides for HLA-A2. In contrast to the situation in patients with putatively muRas-primed T cells, muRas-LCLs were not able to prime naive T lymphocytes from healthy controls. Vaccination of a pancreatic cancer patient with muRas-LCL induced muRas-specific T cells in PBMCs after 4 weeks. We conclude that genetically modified muRas-LCLs can efficiently present tumor antigens to the immune system and induce antigen-specific cytotoxic T cell responses in vitro and in vivo.

The Clinical Significance of Cytokines and Soluble Forms of Membrane-Derived Activation Antigens in the Serum of Patients with Hodgkin's Disease

In a search for specific serum markers with prognostic impact in Hodgkins Disease (HD), we evaluated the clinical significance of several cytokines (IL-1 beta, IL-2, IL-3, IL-6, G-CSF, GM-CSF, TNF-alpha) and soluble forms of membrane-derived antigens (sCD4, sCD8, sCD23, sCD25, sCD30) in the serum of patients with untreated HD. Elevations of three groups of serum factors were observed: Firstly, elevations of the hematopoietic cytokines GM-CSF (detected in 39%), IL-6 (57%) and IL-3 (13%), which occurred simultaneously in the majority of the cases; secondly, simultaneous elevations of the inflammatory cytokines TNF-alpha and IL-1 beta (detected in 7%); and finally, elevations of membrane-derived activation antigens sCD8, sCD25, and sCD30. While the cytokine levels did not correlate with other obvious parameters, the membrane-derived activation antigens sCD8, sCD25 and sCD30 were associated with a poor prognosis. Only sCD30 correlated with disease activity and holds promise for the follow-up of patients in remission. Further investigations of these parameters at the cellular level might help to elucidate the enigmatic biology of HD.

Dose-escalated CHOEP for the treatment of young patients with aggressive non-Hodgkin's lymphoma: I. A randomized dose escalation and feasibility study with bi- and tri-weekly regimens

BACKGROUND To determine the maximum tolerated dose of a bi- and tri-weekly combination chemotherapy with cyclophosphamide, doxorubicin, vincristine and prednisone plus etoposide (CHOEP) regimen without stem-cell support. PATIENTS AND METHODS Randomized phase I/II multicenter four-level (cyclophosphamide: 1000-1200-1400-1600 mg/m2; doxorubicin: 55-60-65-70 mg/m2; etoposide: 375-450-525-600 mg/m2) dose escalation study with CHOEP-14 and CHOEP-21 in young patients (18-60 years) with newly diagnosed aggressive non-Hodgkins lymphoma. Dose-limiting toxicity was defined as thrombocytopenia <80,000/mm3 and leukocytopenia <2500/mm3 on days 16 (CHOEP-14) and 23 (CHOEP-21) or prolonged (>4 days) leukocytopenia (<1000/mm3) or thrombocytopenia (<20,000/mm3). RESULTS One hundred and thirty-nine patients (high-CHOEP-14: 47, high-CHOEP-21: 92) were randomly allocated to the study. Maximal tolerated dose was level 2 for CHOEP-14 and level 4 for CHOEP-21. With a less favorable profile of patients in CHOEP-14, 4-year event-free survival was 47.9% after high-CHOEP-14 and 66.2% after high-CHOEP-21, 4-year overall survival 62.1% after high-CHOEP-14 and 73.4% after high-CHOEP-21, respectively. CONCLUSION Significant dose escalations of CHOEP are possible with granulocyte colony-stimulating factor support, with different chemotherapy models favoring the maximally escalated bi- or tri-weekly regimen, respectively. Because a higher total dose can be achieved with six cycles of the tri-weekly compared with the biweekly regimen, CHOEP-21 at dose escalation level 3 was chosen for a nationwide randomized comparison with baseline CHOEP-21 in a subsequent phase III trial.

Gene-modified spontaneous Epstein-Barr virus-transformed lymphoblastoid cell lines as autologous cancer vaccines: mutated p21 ras oncogene as a model.

The transfer of genes coding for tumor antigens (Ags) into Ag-presenting cells is a promising strategy to develop cancer vaccines for patients not limited by major histocompatibility complex restriction. To test whether autologous lymphoblastoid cell lines (LCL) can be used as an alternative to dendritic cells for Ag presentation, we established LCL by spontaneous growth in cyclosporin-containing medium in vitro (SP-LCL). SP-LCL, which have an advantage over B95-8-transfected LCL in that they carry no risk of introducing a new infectious agent when used for vaccination, served as a permanent source for transfection with an episomal Epstein-Barr virus-based expression vector encoding the Ki-ras p21 oncogene carrying a point mutation at codon 12 (muRas) as a model tumor Ag. The ability of muRas-transfected SP-LCL (muRas-LCL) to induce immunoreactivity was determined in vitro. After electroporation with an optimized protocol, muRas-LCL expressed mutated ras peptides for a considerable period of time (at least 8 weeks) on the cell surface. The transfection procedure did not affect the expression of the costimulatory molecules B7.1, intercellular adhesion molecule-1, and leukocyte function associated Ag-3 by SP-LCL. muRas-LCL were able to induce muRas-specific cytotoxic T lymphocytes from three of three healthy donors and one of one patient with pancreatic carcinoma. Our results suggest that the gene transfer of muRas sequences to SP-LCL leads to an endogenous processing of this Ag in the major histocompatibility complex class I pathway and to functional presentation of antigenic peptides to CD8+ T lymphocytes. Autologous SP-LCL can serve as an unlimited renewable source for autologous cellular vaccines and appear to be good candidates as presenters for a wide range of human tumor Ags.

Advances in molecular biology diagnostic and treatment of B-cell malignancies

While some improvement was achieved by the addition of etoposide and shortening the treatment intervals from 3 to 2 weeks (CHOEP-14), the best results in young, good-prognosis patients [age-adjusted (aa) International Prognostic Index (IPI) 0, 1] have been achieved with six cycles of a CHOP-like chemotherapy in combination with the anti-CD20 antibody rituximab. The role of additional radiotherapy in this setting remains to be determined. With this approach, 2-year event-free survival rates of >90% and overall survival of >95% can be achieved in a very favorable subgroup (patients without IPI risk factor and no bulky disease), while further improvement is warranted for the less favorable subgroup (event-free survival only 77%). For young poor-prognosis patients (aaIPI >_2), the 5-year survival is 50%, and progress has not been convincingly and specifically demonstrated in these patients. Ongoing studies will show whether dose-dense conventional or high-dose chemotherapy regimens requiring stem cell support in combination with rituximab will result in similar improvements of outcome, as has been reported recently for young patients with good-prognosis aggressive lymphoma. In elderly patients, interval reduction of CHOP from 3 to 2 weeks (CHOP-14) and addition of rituximab to CHOP-21 achieved similar improvements in outcome. The ongoing RICOVER-60 trial of the DSHNHL evaluates whether the combination of the two approaches (R-CHOP-14) can further improve the prognosis of elderly patients. Gene analysis of tumor burden may indicate patient prognosis more precisely than the IPI score, but prospective trials are required to find the optimal treatment for each patient. Annals of Oncology 16 (Supplement 2): ii105, 2005 doi:10.1093/annonc/mdi720