Antibody therapy of lymphoma

The availability of rituximab and the possible imminent availability of two new radiolabelled monoclonal anti-CD20 antibodies (Yttrium-90 (90Y)-ibritumomab and Iodine-131(131I)-tositumomab) have captured much attention in the treatment of lymphoma. The chimeric monoclonal anti-CD20 antibody, rituximab has truly heralded a new era for the treatment of lymphoma and human malignancies. The full potential of antibody-based therapy to improve the outcome in patients with B-cell non-Hodgkin's lymphoma has yet to be defined, but recent data suggests that the combination of chemotherapy plus rituximab may significantly improve outcome for patients with aggressive lymphoma over chemotherapy alone. Highly promising data are also emerging for the use of rituximab in combination with chemotherapy in other types of lymphoma. New advances in antibody therapy, driven by new technologies and defining novel antigen targets, offer the promise of more effective tumour specific therapies. Combinations of antibodies, either conjugated with radioisotopes or unlabelled, used with chemotherapy are likely to provide definitive advances in the treatment of lymphoma in the immediate future.     

Rituximab and other emerging monoclonal antibody therapies for lymphoma

The recent approval of rituximab, gemtuzumab ozogamicin, alemtuzumab and ibritumomab tiuxetan by the FDA in the US revealed clear evidence that monoclonal antibodies (mAbs) have significant roles in the current treatment of haematologic malignancies. Among the mAbs under clinical development, anti-CD20 mAbs have been most extensively investigated and have shown definitive clinical efficacy. Rituximab is a genetically engineered chimeric anti-CD20 mAb, with mouse variable and human constant regions. Consecutive clinical trials conducted in the US, Europe and Japan have revealed that rituximab is a highly effective agent with acceptable toxicities against indolent and aggressive B cell non-Hodgkin’s lymphomas (B-NHLs) as a single agent and in combination with cytotoxic drugs. A recent French Phase III study in elderly patients with untreated aggressive B-NHL suggested that the addition of rituximab to standard CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone) chemotherapy increases the complete response rate and prolongs event-free and overall survival. Lymphoma cells are inherently sensitive to radiation. The aim of radioimmunotherapy is to use the mAb to target radiation to lymphoma tissue while minimising toxicity to normal cells. The clinical trials of ⁹⁰Y ibritumomab tiuxetan and ¹³¹I tositumomab showed they have definitive efficacy in relapsed indolent B-NHL with acceptable toxicities. A recent comparative study in relapsed indolent B-NHL showed that ⁹⁰Y ibritumomab tiuxetan produces higher response rates than rituximab. In addition, BL22, a recombinant anti-CD22 immunotoxin, showed significant efficacy in patients with chemotherapy-resistant hairy cell leukaemia. MAbs will have significant roles in the treatment of lymphoid malignancies in the future.     

Monoclonal antibodies in the treatment of non-Hodgkin's lymphoma

Antibody-based therapeutic approaches have had a significant impact in the treatment of non-Hodgkin's lymphoma (NHL). Rituximab's development as an anti-CD20 antibody heralded a new era in treatment approaches for NHL. While rituximab was first shown to be effective in the treatment of relapsed follicular lymphoma, it is now standard monotherapy for front-line treatment of follicular lymphoma, and is also used in conjunction with chemotherapy for other indolent, intermediate and aggressive B-cell lymphomas. The development of rituximab has led to intense interest in this type of therapeutic approach and to development and approval of the radioimmunoconjugates of rituximab, (90)Y-ibritumomab tiuxetan and (131)I-tositumomab, which have added to the repertoire of treatments for relapsed follicular lymphoma and increased interest in developing other conjugated antibodies. Since rituximab is a chimeric antibody, there is a need to develop fully humanised antibodies, such as IMMU-106 (hA20), in order to minimise infusion reactions and eliminate the development of human antibodies against the drug. Further clinical evaluation of antibodies has been based largely on our knowledge of antigen expression on the surface of lymphoma cells and has led to the development of antibodies against CD22 (unconjugated epratuzumab and calicheamicin conjugated CMC-544 [inotuzumab ozogamicin]), CD80 (galiximab), CD52 (alemtuzumab), CD2 (MEDI-507 [siplizumab]), CD30 (SGN-30 and MDX-060 [iratumumab]), and CD40 (SGN-40). Furthermore, the VEGF (vascular endothelial growth factor) inhibitor bevacizumab, which was first approved for the treatment of colon cancer is currently under investigation in NHL, and agonists rather than antibodies to TRAIL (tumour necrosis factor-related apoptosis-inducing ligand) [rApo2L/TRAIL, HGS-ETR1{mapatumumab}, HGS-ETR2] are currently being investigated as treatments for both advanced solid tumours and NHL. Knowledge of the ability of cancer cells to become resistant to a targeted therapy by activating an alternative pathway to evade apoptosis has driven studies that combine antibodies such as epratuzumab plus rituximab (ER) or ER plus chemotherapy with CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) [ER-CHOP], inotuzumab ozogamicin plus rituximab, alemtuzumab plus CHOP (CHOP-C), bevacizumab plus rituximab, and now the combination of rApo2L/TRAIL plus rituximab. As a result of the expansion of research in this area, several treatment-specific adverse effects have been noted, including infusion-related reactions for rituximab, myelosuppression secondary to (90)Y-ibritumomab tiuxetan and (131)I-tositumomab, and immunosuppression leading to infectious complications for alemtuzumab. Also, soluble forms of the antigens (sCD30) are now being investigated as potential mechanisms of resistance to antibody treatments by binding the antibody before the drug can bind to the lymphoma cell. In addition, it has also become apparent that these antibodies often have a dose-dependent half-life (rituximab) or long half-lives of up to 2-3 weeks (epratuzumab and galiximab) with a consequent delay to a response, thus influencing how long we should wait for a response before declaring an antibody to be ineffective. Antibody-based therapeutic approaches have already had a profound impact on the treatment of NHL, and it is almost certain that, as their clinical development progresses, we will continue to refine the optimum methods of incorporating these drugs in NHL treatment in order to offer our patients the best clinical benefits.     

Treatment of acute leukaemias with monoclonal antibodies: current status and future prospects

Monoclonal antibodies are a new class of agents targeted at specific receptors on cancer cells. In addition to direct cellular effects, antibodies can carry substances to the targeted cells, such as radioactive isotopes, toxins, and antineoplastic agents. At present monoclonal antibodies, directed against both lymphoid antigens (CD 20 and CD 52) and a myeloid antigen (CD33) are available for clinical use. In ALL, rituximab, a humanized anti CD20 antibody, has been combined to chemotherapy mainly in mature B-ALL and Burkitt's lymphoma and preliminary results are promising. Alemtuzumab is an anti-CD52 humanized antibody, which showed anti-tumour activity in CLL; clinical effects were observed in some patients with relapsed adult ALL. Monoclonal antibodies against myeloid antigens have been prevalently used in acute myeloid leukaemias (AML), where the most utilised immunological target is CD33. The CD33 molecule is expressed by approximately 90% of AMLs but not on CD34(+) bone marrow-resident hematopoietic stem cells. The humanized anti-CD33 monoclonal antibody HuM195 has only modest activity against overt AML, but it can eliminate minimal residual disease. Radioimmunotherapy with beta-particle-emitting isotopes targeting CD33 shows a major efficacy. Targeted chemotherapy with the anti-CD33-calicheamicin construct gemtuzumab ozogamicin (GO) has produced remissions as a single agent in patients with relapsed AML and appears promising when used in combination with standard chemotherapy. GO is composed of a humanized immunoglobulin G(4) (IgG(4)) monoclonal antibody (mAb), targeting the CD33 antigen and linked to a calicheamicin derivative, a cytotoxic anthracycline antibiotic. GO has been approved by FDA as second-line therapy in older patients with AML. The most common adverse effects of monoclonal antibodies are myelosuppression, infusion-related reactions, and hypersensitivity reactions. Rituximab may cause tumour lysis syndrome. Alemtuzumab causes immunosuppression, increasing the risk of infection. GO may cause hepatotoxicity.     

Rituximab: a review of its use in non-Hodgkin's lymphoma and chronic lymphocytic leukaemia

Rituximab (MabThera, Rituxan) is an anti-CD20 monoclonal antibody that induces lysis and apoptosis of normal and malignant human B cells, and sensitises malignant B cells to the cytotoxic effect of chemotherapy. In phase III trials in patients with indolent or aggressive B-cell non-Hodgkin's lymphoma (NHL), intravenous rituximab in combination with chemotherapy was more effective as first- or second-line therapy than chemotherapy alone in providing tumour remission and patient survival. Likewise, in patients with chronic lymphocytic leukaemia (CLL), rituximab in combination with chemotherapy appeared more effective than chemotherapy alone as either first- or second-line treatment. In addition, rituximab maintenance therapy was shown to significantly prolong tumour remission and patient survival in patients with indolent B-cell NHL or CLL. The combination of rituximab with cyclophosphamide, doxorubicin, vincristine and prednisone (CHOP) was cost effective as first-line therapy for advanced-stage diffuse large B-cell NHL compared with CHOP alone. Rituximab, either alone or in combination with chemotherapy, was generally well tolerated in patients with NHL or CLL. Overall, rituximab in combination with chemotherapy, is a valuable option for first- and second-line therapy in patients with advanced-stage indolent or aggressive B-cell NHL, and possibly those with B-cell CLL, and is included in current treatment guidelines for these indications. The drug is also potentially useful as maintenance therapy in patients with indolent B-cell NHL or CLL.     

Treatment of systemic lupus erythematosus with epratuzumab

Systemic lupus erythematosus is a prototypic autoimmune disease characterized by abnormalities in the activity of B-cells and T-cells. A novel specific treatment for autoimmune diseases is B-cell depletion with monoclonal antibodies. Epratuzumab is a monoclonal antibody that targets CD22 antigen on B-cells. Initial phase II and two terminated early phase III studies suggest that treatment of systemic lupus erythematosus with this immunomodulatory agent is effective, well tolerated and significantly improves the patient's quality of life. In vitro studies and clinical trials with non-Hodgkin lymphoma patients indicate epratuzumab can potentially serve as a complementary drug in combination therapy with another inhibitor of B-cell activity, rituximab, which is a monoclonal anti-CD20 antibody.     

利妥昔单抗治疗恶性淋巴瘤的进展

利妥昔单抗（人源化CD20单克隆抗体，商品名美罗华）是首个批准用于治疗表达CD20恶性淋巴省的单克隆抗体，广泛应用于低度恶性非雷奇金琳巴瘤（NHL）、侵袭性NHL，亦试用于霍奇金淋巴瘤（HL）及其他B细胞性恶性肿瘤。刊蚤昔单抗联合细胞毒性药物的疗效已在B细胞NHL的有关临床试验中得到证实。目前该药已被批准用于侵袭性淋巴瘤和惰性淋巴瘤的一线治疗，其维持治疗滤泡性淋巴瘤亦得出鼓舞人心的结果．利妥昔单抗已成为治疗B细胞性恶性淋巴瘤的重要手段之一。     

Rituximab and other emerging monoclonal antibody therapies for lymphoma

The recent approval of rituximab, gemtuzumab ozogamicin, alemtuzumab and ibritumomab tiuxetan by the FDA in the US revealed clear evidence that monoclonal antibodies (mAbs) have significant roles in the current treatment of haematologic malignancies. Among the mAbs under clinical development, anti-CD20 mAbs have been most extensively investigated and have shown definitive clinical efficacy. Rituximab is a genetically engineered chimeric anti-CD20 mAb, with mouse variable and human constant regions. Consecutive clinical trials conducted in the US, Europe and Japan have revealed that rituximab is a highly effective agent with acceptable toxicities against indolent and aggressive B cell non-Hodgkin's lymphomas (B-NHLs) as a single agent and in combination with cytotoxic drugs. A recent French Phase III study in elderly patients with untreated aggressive B-NHL suggested that the addition of rituximab to standard CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone) chemotherapy increases the complete response rate and prolongs event-free and overall survival. Lymphoma cells are inherently sensitive to radiation. The aim of radioimmunotherapy is to use the mAb to target radiation to lymphoma tissue while minimising toxicity to normal cells. The clinical trials of 90Y ibritumomab tiuxetan and (131)I tositumomab showed they have definitive efficacy in relapsed indolent B-NHL with acceptable toxicities. A recent comparative study in relapsed indolent B-NHL showed that 90Y ibritumomab tiuxetan produces higher response rates than rituximab. In addition, BL22, a recombinant anti-CD22 immunotoxin, showed significant efficacy in patients with chemotherapy-resistant hairy cell leukaemia. MAbs will have significant roles in the treatment of lymphoid malignancies in the future.     

Bexxar (Corixa/GlaxoSmithKline)

Bexxar (131I tositumomab) is a radiolabeled anti-CD20 monoclonal antibody for the treatment of relapsed and refractory follicular/low-grade and transformed non-Hodgkin's lymphoma. It has shown high response rates with durable complete remissions in patients who have received either prior chemotherapy or rituximab (Rituxan, MabThera; IDEC Pharmaceuticals Corp/Genentech/F Hoffmann La Roche). Complications include myelosuppression, secondary acute leukemia, myelodysplasia and hypothyroidism. The role of this promising new agent is being defined in phase II and III trials. In February 2001, ABN Amro Predicted launch in 2001 and sales of US $25 million rising to US $70 million in 2003 [422363]. Corixa and GlaxoSmithKline anticipate a launch in the US in 2002 [424619].     

Rituximab: as first-line maintenance therapy following rituximab-containing therapy for follicular lymphoma

Rituximab is a recombinant chimeric murine/human monoclonal IgG(1-κ) antibody. It binds specifically to the CD20 antigen on normal and malignant B lymphocytes and produces complement-dependent and antibody-dependent cytotoxicity and induces apoptosis in these cells. Prolonged treatment with rituximab in patients with follicular lymphoma results in a sustained reduction in circulating B lymphocytes. Two years of single-agent maintenance therapy with rituximab significantly prolonged progression-free survival (primary endpoint) compared with observation in patients with follicular lymphoma who were responsive to first-line induction therapy with rituximab plus chemotherapy. Furthermore, maintenance therapy with rituximab significantly delayed the time to the next antilymphoma treatment and the next chemotherapy compared with observation in these patients. Rituximab had an acceptable tolerability profile as single-agent maintenance therapy in patients with follicular lymphoma with no new or unexpected adverse events compared with induction therapy.     

Mechanism of action of rituximab

Rituximab, the humanized chimeric anti-CD20 monoclonal antibody, represents a powerful tool for treating B-cell malignancies and is licensed for the treatment of relapsed or chemorefractory low-grade or follicular non-Hodgkin's lymphoma (NHL). It has a unique mode of action and can induce killing of CD20+ cells via multiple mechanisms. The direct effects of rituximab include complement-mediated cytotoxicity and antibody-dependent cell-mediated cytotoxicity, and the indirect effects include structural changes, apoptosis, and sensitization of cancer cells to chemotherapy. In vitro studies have made a significant contribution to the understanding of these mechanisms of action and have led to the development of innovative and effective treatment strategies to optimize patient response. The most significant of these strategies is the combination of rituximab and CHOP chemotherapy (cyclophosphamide, doxorubicin, vincristine and prednisone), which is proving a highly effective combination in the treatment of NHL. However, all patients do not respond equally well to rituximab, and in vitro studies have identified a possible mechanism of resistance involving the anti-complement inhibitors CD55 and CD59. Neutralizing antibodies to CD55 and CD59 can overcome resistance to rituximab-mediated complement-mediated cytotoxicity in vitro. This paper overviews our understanding of the mechanisms of action of rituximab and identifies how this knowledge could be applied in a clinical setting to maximize response in both sensitive and resistant patients.     

Ofatumumab,a second-generation anti-CD20 monoclonal antibody,for the treatment of lymphoproliferative and autoimmune disorders

Background: Lymphoproliferative and autoimmune disorders share monoclonal dysregulation and survival advantage of B-lymphocytes. Thus, therapies directed towards eliminating B-cells will play an important role as CD20 is exclusively expressed in B-lymphocytes and its modulation by monoclonal antibodies such as rituximab has improved outcomes in lymphoproliferative and autoimmune disorders. Ofatumumab is a new, fully human anti-CD20 antibody and has been shown to be effective and safe, but its role in these conditions is still unclear. Objectives: To describe the preclinical and clinical data available on ofatumumab for the treatment of lymphoproliferative and autoimmune disorders. Methods: An extensive search of published articles and abstracts on preclinical and clinical studies with ofatumumab was undertaken. Conclusions: Ofatumumab is a second-generation anti-CD20 antibody that has been demonstrated to be safe and efficacious in patients with lymphoproliferative and autoimmune disorders. Ofatumumab is fully human, attaches to a newly identified epitope and shows lower off-rates and improved complement-dependent cytotoxicity. Initial data present ofatumumab as an attractive agent with lower rates of infusion-related events than rituximab. Ongoing Phase III trials in patients with follicular lymphoma, chronic lymphocytic leukemia and rheumatoid arthritis are ongoing, and Phase II trials in patients with aggressive lymphoma and multiple sclerosis are also under development.     

Aggressive lymphoma: improving treatment outcome with rituximab

The standard therapy for patients with aggressive lymphoma is cyclophosphamide, doxorubicin, vincristine and prednisone (CHOP) chemotherapy, which achieves a complete response in more than 60% of patients but is curative in only about 40-50%. More aggressive and/or dose-intensified chemotherapy regimens have failed to provide significant survival advantages compared with CHOP, and may have higher toxicity. Rituximab, a chimeric monoclonal antibody to the CD20 antigen, is effective as monotherapy in aggressive lymphoma and in combination with chemotherapy has demonstrated high response rates in phase II trials. A scheduled interim analysis of a randomized, prospective trial comparing rituximab plus CHOP with CHOP alone in elderly patients with untreated diffuse large B-cell lymphoma has shown significantly better response rates and survival with rituximab plus CHOP compared with CHOP alone. These results represent the first significant improvement in overall survival over CHOP in aggressive lymphoma for over 20 years. The addition of rituximab was not associated with significant additional toxicity over that seen with CHOP alone. Ongoing studies are underway to establish whether the survival benefit of rituximab plus CHOP is seen in younger patient populations. Rituximab in combination with chemotherapy is also being evaluated as salvage treatment for patients who relapse after initial chemotherapy. In a preliminary analysis of a study in 50 patients with refractory or relapsed aggressive lymphoma, rituximab plus etoposide, prednisone, vincristine, cyclophosphamide and doxorubicin (EPOCH) chemotherapy has demonstrated promising results when used as sole salvage therapy and as an induction therapy prior to autologous stem-cell transplantation, again without significant additional toxicity.     

Ten years of rituximab in NHL

Background: Rituximab, a chimeric mouse/human monoclonal antibody targeting the pan-B-cell antigenic marker CD20, was the first monoclonal antibody licensed for use in the treatment of cancer. Objective: This review focuses on the impact of rituximab in the treatment of patients with B-cell non-Hodgkin lymphoma (NHL). Methods: Three key areas related to the use of rituximab in B-cell NHL are discussed: mechanism of action, clinical efficacy in both indolent and aggressive disease, and safety of its use as both monotherapy and in combination with chemotherapy. Results/conclusions: Rituximab has demonstrated significant clinical efficacy in the treatment of NHL, particularly in combination with chemotherapy, and its use has revolutionized the treatment of both indolent and aggressive B-cell NHL over the past decade. Furthermore, consistent toxicity data have been obtained with a safe and tolerable profile in most patients.     

Ibritumomab: new drug. Interesting concept, disappointing in practice

(1) First-line standard chemotherapy for follicular non-Hodgkin's lymphoma is based on alkylating agents. For patients who relapse, rituximab, a monoclonal antibody targeting CD20 receptors on tumour cells, induces remissions lasting at least 9 months in about 50% of cases. However, the infusions are often poorly tolerated and reactions can be severe. (2) Ibritumomab is a monoclonal antibody similar to rituximab. It is not in itself cytotoxic. Yttrium-90-radiolabeled ibritumomab has been licensed in the EU since 2004 for local tumour radiotherapy, administered shortly after rituximab therapy. It is approved as a third-line treatment for follicular lymphoma, after rituximab failure. (3) The only available comparative trial, lasting 4 years, involved 143 patients who were treated either with rituximab followed by radiolabeled ibritumomab, or with rituximab alone. The sequential treatment yielded more tumour responses but the time to lymphoma progression and clinical outcomes did not differ between the two treatment groups. (4) In a non comparative trial about half the patients in whom rituximab had failed responded to rituximab + radiolabeled ibritumomab, but lymphoma progression occurred after a median of only 7 months. (5) Patients who receive sequential therapy with rituximab + ibritumomab are exposed to the adverse effects of rituximab plus supplementary gastrointestinal, respiratory, haematological and infectious risks, mainly due to persistent myelosuppression. (6) The radiation hazard for the patient (irradiation of healthy organs) and for the environment appears to be limited, because of the short reach of beta particle emission (5 mm) and the short half-life of yttrium 90 (2.7 days). (7) Yttrium 90 radiolabeling requires special equipment and specialised personnel working in an approved centre, which can restrict access to treatment. (8) Radiolabeled ibritumomab appears to have a negative risk-benefit balance, and should only be used in clinical trials.     

Current concepts and future directions in radioimmunotherapy

Radioimmunotherapy relies on the principles of immunotherapy, but expands the cytotoxic effects of the antibody by complexing it with a radiation-emitting particle. If we consider radioimmunotherapy as a step beyond immunotherapy of cancer, the step was prompted by the (relative) failure of the latter. The conventional way to explain the failure is a lack of intrinsic killing effect and a lack of penetration into poorly vascularized tumor masses. The addition of a radioactive label (usually a β-emitter) to the antibody would improve both. Radiation is lethal and the type of radiation used (beta rays) has a sufficient range to overcome the lack of antibody penetration. At present, the most successful (and FDA approved) radioimmunotherapy agents for lymphomas are anti-CD20 monoclonal antibodies. Rituximab (Rituxan(?)) is a chimeric antibody, used as a non-radioactive antibody and to pre-load the patient when Zevalin(?) is used. Zevalin(?) is the Yttrium-90 ((90)Y) or Indium-111 ((111)In) labeled form of Ibritumomab Tiuxetan. Bexxar(?) is the Iodine-131 ((131)I) labeled form of Tositumomab. Ibritumomab Tiuxetan and Tositumomab are murine anti-CD20 monoclonal antibodies, not chimeric antibodies. Promising research is being done to utilize radioimmunotherapy earlier in the treatment algorithm for lymphoma, including as initial, consolidation, and salvage therapies. However, despite more than 8 years since initial regulatory approval, radioimmunotherapy still has not achieved widespread use due to a combination of medical, scientific, logistic, and financial barriers. Other experimental uses for radioimmunotherapy include other solid tumors to treat infections. Optimization can potentially be done with pre-targeting and bi-specific antibodies. Alpha particle and Auger electron emitters show promise as future radioimmunotherapy agents but are mostly still in pre-clinical stages.     

Rituximab therapy for indolent non-Hodgkin's lymphoma

Indolent non-Hodgkin's lymphomas (NHLs) are essentially incurable with current treatments. Rituximab is a specific anti-CD20 chimeric monoclonal antibody against the CD20 antigen, which is stably expressed on most B-cells (from the pre-B-cell stage). Compared with chemotherapy, rituximab has an excellent tolerability profile, making it a good therapeutic option for patients with indolent NHL. In the pivotal study for rituximab, patients with relapsed or refractory indolent or follicular lymphoma (FL) had an overall response rate of 50%. There is evidence that first-line rituximab therapy may be associated with better response rates; in previously untreated FL with a low tumor burden, rituximab monotherapy has produced an overall response rate of 73%. Attempts to improve response rates to rituximab by increasing the dose or frequency of dosing showed that the addition of four extra infusions of rituximab (in addition to the standard treatment schedule) resulted in an overall response rate of 76% in patients with FL. Augmenting rituximab with cytokines is also an option for increasing response rates in patients with indolent NHL. In a trial by the Nordic Lymphoma Study Group in patients with previously untreated or first-relapse indolent NHL, who had stable disease or a partial response after four doses of rituximab, 48% of the patients treated with rituximab plus interferon-alpha2a achieved a complete response. A further option is to combine rituximab with chemotherapy. Interim analyses from the East German Study Group have shown that rituximab plus mitoxantrone, chlorambucil and prednisolone (MCP) resulted in overall response rates of 89% in patients with untreated indolent lymphoma. Rituximab is therefore an excellent treatment option both as first-line and as salvage therapy for patients with indolent NHL.     

Human therapeutic antibodies.

The development of genetic engineering technologies has today advanced to the point where the generation of high-affinity human antibodies against therapeutic targets is not a major hurdle. Rather, it is the selection of target molecules in, for example, cancer therapy that poses a challenge. Targets that are not merely passive acceptors but those that signal into the cell are preferred. Recent advances in the clinical use of antibody-based therapy--such as anti-CD20 (rituximab) for the treatment of non-Hodgkin's lymphoma and anti-tumour-necrosis-factor-alpha for Crohn's disease--as well as novel antibody designs and improved understanding of the mode of action of current antibodies lend great hope to the future of this therapeutic approach.     

CD20-targeting in B-cell malignancies: novel prospects for antibodies and combination therapies

Expression of CD20 antigen by the most of transformed B cells is believed to be the driving force for targeting this molecule by using anti-CD20 monoclonal antibodies. While it is true that most lymphoma/leukemia patients can be cured, these regimens are limited by the emergence of treatment resistance. Based on these observations, development of anti-CD20 monoclonal antibodies and combination therapies have been recently proposed, in particular with the aim to optimize the cytotoxic activity. Here we outline a range of new experimental agents concerning the CD20 positive B-cell tumors which provide high benefit from conventional therapy.     

Construction and characterization of monoclonal antibodies against the extracellular domain of B-lymphocyte antigen CD20 using DNA immunization method

To date, several new anti-CD20 monoclonal antibodies (mAbs) have been developed for potential efficacies compared with familiar mAb rituximab. Despite the recent advances in development of anti-CD20 mAbs for the treatment of B cell malignancies, the efforts should be continued to develop novel antibodies with improved properties. However, the development of mAbs against CD20 as a multi-transmembrane protein is challenging due to the difficulty of providing a lipid environment that can maintain native epitopes. To overcome this limitation, we describe a simple and efficient DNA immunization strategy for the construction of a novel anti-CD20 mAb with improved anti-tumour properties. Using a DNA immunization strategy that includes intradermal (i.d.) immunization with naked plasmid DNA encoding the CD20 gene, we generated the hybridoma cell line D4, which secretes functional mAbs against an extracellular epitope of CD20. Immunocytochemistry analysis and a cell-based enzyme-linked immunosorbent assay using a Burkitt's lymphoma cell line showed that D4 mAbs are capable of binding to native extracellular epitopes of CD20. Moreover, the binding specificity of D4 mAbs was determined by western blot analysis. Cell proliferation was examined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis was detected by the annexin V/propidium iodide staining and dye exclusion assay. The results showed that D4 anti-CD20 mAbs produced by DNA immunization exhibit potent growth inhibitory activity and have superior direct B-cell cytotoxicity compared to rituximab. We propose that antibody-induced apoptosis is one of the mechanisms of cell growth inhibition. Taken together, the data reported here open the path to DNA-based immunization for generating pharmacologically active monoclonal antibodies against CD20. In addition, the data support future in vivo animal testing and subsequent procedures to produce a potential therapeutic mAb.