Development of antibodies for cancer therapy

Antibody therapies have become an important component in the management of malignant disease. Recombinant technology offers enormous opportunities to tailor antibodies to meet clinical requirements. This includes the reduction of immunogenicity and the development of smaller antibody fragments that can be incorporated into fusion proteins. Antibodies can block tumour growth factors or their receptors, activate immunological attack on the tumour, or be used to deliver payloads such as radioisotopes, cytotoxic drugs or toxins. Pretargeting includes streptavidin/biotin systems and antibody-directed enzyme prodrug therapy (ADEPT). ADEPT uses an antibody–enzyme complex to deliver a prodrug-activating enzyme to tumours for selective prodrug conversion at the tumour site. New antibody targets, refined antibodies, antibody fusion proteins, combination therapies and the use of antibodies as adjuvant therapy are important topics in the development of antibody therapy against cancer.     

Development of antibodies for cancer therapy

Antibody therapies have become an important component in the management of malignant disease. Recombinant technology offers enormous opportunities to tailor antibodies to meet clinical requirements. This includes the reduction of immunogenicity and the development of smaller antibody fragments that can be incorporated into fusion proteins. Antibodies can block tumour growth factors or their receptors, activate immunological attack on the tumour, or be used to deliver payloads such as radioisotopes, cytotoxic drugs or toxins. Pretargeting includes streptavidin/biotin systems and antibody-directed enzyme prodrug therapy (ADEPT). ADEPT uses an antibody-enzyme complex to deliver a prodrug-activating enzyme to tumours for selective prodrug conversion at the tumour site. New antibody targets, refined antibodies, antibody fusion proteins, combination therapies and the use of antibodies as adjuvant therapy are important topics in the development of antibody therapy against cancer.     

Monoclonal antibodies for brain tumour treatment

Conventional treatment of brain tumours includes surgical, radiotherapeutic and chemotherapeutic modalities. Nonetheless, the outcome of patients with brain tumours, in particular glioblastoma, remains poor. Immunotherapy with armed or unarmed monoclonal antibodies targeting tumour-specific antigens has emerged in the last two decades as a novel potential adjuvant treatment for all types of neoplasia. Many challenges to its implementation as a safe and viable therapy for brain tumours still need to be addressed; nevertheless, results from ongoing Phase I/II clinical trials are encouraging, as disease stabilisation and patient survival prolongation have been observed. Advances in preclinical and clinical research indicate that treatment of brain tumours with monoclonal antibodies can be increasingly adjusted to the characteristics of the targeted tumour and its environment. This aspect relies on the careful selection of the target antigen and corresponding specific monoclonal antibody, and antibody format (size, class, affinity), conjugation to the appropriate toxin or radioactive isotope (half-life, range), and proper compartmental administration.     

Monoclonal antibodies for brain tumour treatment

Conventional treatment of brain tumours includes surgical, radiotherapeutic and chemotherapeutic modalities. Nonetheless, the outcome of patients with brain tumours, in particular glioblastoma, remains poor. Immunotherapy with armed or unarmed monoclonal antibodies targeting tumour-specific antigens has emerged in the last two decades as a novel potential adjuvant treatment for all types of neoplasia. Many challenges to its implementation as a safe and viable therapy for brain tumours still need to be addressed; nevertheless, results from ongoing Phase I/II clinical trials are encouraging, as disease stabilisation and patient survival prolongation have been observed. Advances in preclinical and clinical research indicate that treatment of brain tumours with monoclonal antibodies can be increasingly adjusted to the characteristics of the targeted tumour and its environment. This aspect relies on the careful selection of the target antigen and corresponding specific monoclonal antibody, and antibody format (size, class, affinity), conjugation to the appropriate toxin or radioactive isotope (half-life, range), and proper compartmental administration.     

Anti-idiotype antibody vaccine therapy for cancer

The use of anti-idiotype (Id) antibodies as vaccines to stimulate antitumour immunity is one of several promising immunologic approaches to the therapy of cancer. Extensive studies in animal tumour models have demonstrated the efficacy of anti-Id vaccines in preventing tumour growth and curing mice with established tumours. A number of monoclonal anti-Id antibodies that mimic distinct human tumour-associated antigens (TAAs) have been developed and tested in the clinic, and demonstrate encouraging results. In general, the antigen mimicry by anti-Id antibodies has reflected structural homology in the majority of the cases, and amino acid sequence homology in a few of them. The greatest challenge of immunotherapy by means of anti-Id vaccines is to identify the optimal anti-Id antibody that will function as a true surrogate antigen for a TAA system, and ideally will generate both humoral and cellular immune responses. Although several clinical studies have shown enhanced survival of patients receiving anti-Id vaccines, the efficacy of these vaccines will depend on the results of several randomised Phase III clinical trials that are currently planned or ongoing.     

Anti-idiotype antibody vaccine therapy for cancer

The use of anti-idiotype (Id) antibodies as vaccines to stimulate antitumour immunity is one of several promising immunologic approaches to the therapy of cancer. Extensive studies in animal tumour models have demonstrated the efficacy of anti-Id vaccines in preventing tumour growth and curing mice with established tumours. A number of monoclonal anti-Id antibodies that mimic distinct human tumour-associated antigens (TAAs) have been developed and tested in the clinic, and demonstrate encouraging results. In general, the antigen mimicry by anti-Id antibodies has reflected structural homology in the majority of the cases, and amino acid sequence homology in a few of them. The greatest challenge of immunotherapy by means of anti-Id vaccines is to identify the optimal anti-Id antibody that will function as a true surrogate antigen for a TAA system, and ideally will generate both humoral and cellular immune responses. Although several clinical studies have shown enhanced survival of patients receiving anti-Id vaccines, the efficacy of these vaccines will depend on the results of several randomised Phase III clinical trials that are currently planned or ongoing.     

Therapeutic milestones in tumor therapy

The treatment of cancer has witnessed dramatic improvement throughout the 20th century. All three classical pillars of cancer therapy, tumour surgery, radiation oncology and medical oncology have provided impressive results which early in our century had not been anticipated to this extent. Particular mile stones of treatment include the discovery that prostate cancer is a hormone-sensitive tumour and the development of curative chemotherapy for many haematological neoplasms, or advanced testicular cancer. Adjuvant systemic treatment has led to significant improvement in the prognosis of patients with breast cancer and other solid tumours. More recently preoperative chemotherapy is gaining an important role in particular disorders such as non-small cell lung cancer. The most fascinating advances, however, have originated from our improved understanding of the molecular and cellular basis of malignancy. The development of monoclonal antibodies targeting specific tumour antigens, tumour vaccines and cancer gene therapy are all clinically relevant results of diligent basic and translational experimental cancer research.     

Limited improvement of tumour diagnosis by the simultaneous determination of carcinoembryonic antigen (CEA) and of a tumour-associated CEA-related antigen of Mr 128,000 in serum

Two biotin-avidin based enzyme immunoassays were developed using three monoclonal anti-CEA antibodies with distinct epitope- and antigen-specificities. A broadly cross-reactive monoclonal anti-CEA antibody (T84.1) was immobilized on a solid support. Either monoclonal antibody T84.66 or CEA.11 was used as the second, biotin-labeled monoclonal antibody. Both antibodies do not cross-react with normal granulocytes or bile canaliculi. Monoclonal antibody CEA.11 binds to CEA and to an antigen with a relative molecular mass of 128,000 present in extracts from solid carcinomas. Monoclonal antibody T84.66 does not cross-react with the antigen of Mr 128,000. After adsorption of tumour extracts to a CEA-specific immunosorbent, residual activity was measurable by the CEA.11 assay, but not by the T84.66 assay. Serum samples (n = 726) from patients with malignant and non-malignant disease, as well as from healthy volunteers, were analysed by both immunoassays and by two commercial CEA immunoassays. In comparison with the T84.66 assay and the commercial assays, the CEA.11 assay did not significantly increase the sensitivity or specificity of tumour diagnosis.     

Requirements for human antibody cocktails for oncology

Cancer patients receiving antibodies as monotherapy have benefited from these treatments. However, significant improvements can be made that should make the therapy more effective. Applying lessons learned from the natural oligoclonal antibody response that cancer patients mount to their own tumours suggests that cocktails of monoclonal antibodies could be formulated, which may be more effective in treating cancers. The next phase of antibody immunotherapy will include cocktails of monoclonal antibodies. Various requirements for human antibody cocktails are discussed, as well as potential limitations of this approach.     

Serologic and cellular assays to monitor therapy with murine monoclonal antibodies

The emergence of new therapeutic modalities frequently requires development of relevant laboratory assays. This is especially true for the clinical uses of newly developed reagents such as murine monoclonal antibodies. We have described immunofluorescence, immunohistochemical, and enzyme-linked immunosorbent assays (ELISA) for monitoring anticancer therapy with monoclonal antibodies. Immunofluorescence assays performed on single cell suspensions and immunohistochemical assays performed on freshly frozen biopsy tissue have confirmed tissue reactivity of the monoclonal antibody prior to infusion and have been used to measure in vivo antibody binding to target cells as well as antigenic modulation during therapy. An ELISA has been used to measure the serum concentrations of monoclonal antibodies; another ELISA has been used to detect the presence of human antibodies reactive with murine monoclonal antibody. These reproducible assays can be performed with commercially available reagents and provide an important data base for making clinical decisions concerning therapy with monoclonal antibodies.     

Requirements for human antibody cocktails for oncology

Cancer patients receiving antibodies as monotherapy have benefited from these treatments. However, significant improvements can be made that should make the therapy more effective. Applying lessons learned from the natural oligoclonal antibody response that cancer patients mount to their own tumours suggests that cocktails of monoclonal antibodies could be formulated, which may be more effective in treating cancers. The next phase of antibody immunotherapy will include cocktails of monoclonal antibodies. Various requirements for human antibody cocktails are discussed, as well as potential limitations of this approach.     

Monoclonal immunotherapy with human monoclonal antibody(CLN-IgG) in glioma patients

It is well recognized that malignant gliomas escape an immune response by hiding behind the blood-brain barrier and by producing proteins that suppress systemic immunity. However, if gliomas can be made to be more immunogenic or if a tumor vaccine can be produced, then access to all tumor cells including those that infiltrate into the brain can be achieved through the patient's immune response. Several strategies have been investigated for immunotherapy. Laboratory studies and animal models have shown that these immune cells will attack the tumor cell, reduce the size of implanted tumors, and that the immune memory is sufficient to suppress tumor growth when the animal is rechallenges with a tumor implant. Since the development of hybridoma technology, monoclonal antibodies against human cancer cells have been produced and antigens have been identified. Hagiwara reported the production of a human monoclonal antibody, CLN-IgG, made by fusing UC 729-6, human lymphoblastoid B-cell line, with lymphocytes obtained from a patient with the cervical carcinoma. It has been reported that CLN-IgG recognized the antigen expressed in various histological types of human cancers including malignant gliomas. The effect of human monoclonal antibody(CLN-IgG) on malignant brain tumors was evaluated in patients with malignant glioma. Early phase II study was concluded that this specific immunotherapy with CLN-IgG is safe and effective therapy in patients with malignant glioma. We treated 10 cases of malignant gliomas with CLN-IgG. All patients had received radiotherapy and chemotherapy before this immunotherapy using the human monoclonal antibody. The human monoclonal antibody(CLN-IgG) was administered intravenously once or twice/week during 24 weeks. Six cases of glioblastoma, 1 medulloblastoma and 3 cases of potine glioma histologically unverified, were treated. Five cases of 6 glioblastomas died 4 to 12 months after this treatment, 3 cases of pontine glioma showed good responses, 2 cases showed marked decrease of tumor size and 1 case showed no regrowth of tumor on MRI imaging. For the above reasons, Human monoclonal antibody(CLN-IgG) might be useful as an immunotherapy of malignant gliomas.     

13. New biological therapies for breast cancer

The exploitation of biological differences between normal and malignant cells is a logical approach to novel treatments for breast cancer. The potential targets for such therapy include the products of proto-oncogenes and oncogenes, inhibition of growth factor receptor signalling and the immunological exploitation of antigenic differences between normal and malignant cells. Monoclonal antibody technology was heralded as a potential 'magic bullet' for cancer therapy following its discovery in the mid-1970s, but it is only in the past few years that such technology has entered mainstream clinical practice. The humanised murine monoclonal antibody to HER2 (trastuzumab) has significant anti-tumour activity but with minimal toxicity, and has been licensed for use in patients with advanced breast cancer. A different approach has been the use of enzyme inhibitors to interfere with the signalling pathways downstream of growth factor receptors (e.g. farnesyl transferase inhibitors). It is likely that effective targets for such therapies will be identified in the next few years. There have been significant advances in our understanding of human immunology which have coincided with the identification of so-called tumour-associated antigens (TAA). These developments have resulted in a resurgence of interest in tumour immunotherapy. Peptides derived from these TAAs have been used to generate tumour-specific immune responses. An alternative strategy has been to immunise patients using viral vectors and plasmid cDNA encoding the TAA. In some studies, notably those in patients with advanced melanoma, significant clinical responses have been observed. Cell-based strategies including autologous tumour cell vaccines, allogeneic tumour cell vaccines and dendritic cell vaccines have been used, and significant responses have been reported in several studies. Few of these methods have so far been applied to breast cancer, but the possible benefits and drawbacks of such an approach will be discussed.     

Monoclonal antibody therapy for disorders of hemostasis and coagulation

Monoclonal antibody therapies have conducted to not only hematologic malignancies but also disorders of hemostasis and coagulation. This article describes the recent advances of monoclonal antibody therapy for bleeding disorders such as idiopathic thrombocytopenic purpura(ITP), hemophilia A, disseminated intravascular coagulation(DIC), and thrombosis. Rituximab, chimeric anti-CD20 monoclonal antibody treatment has a valuable effect in the patients with ITP, and clinical trials using anti-CD40 ligand monoclonal antibody for ITP are underway. Anti-CD40 ligand monoclonal antibody can be an alternative therapy for hemophilia A patients with inhibitors to factor VIII. In thrombosis, anti-tissue factor monoclonal antibody and anti-factor IX(a) monoclonal antibody were established as novel anticoagulant regents. Plasminogen activator inhibitor-1(PAI-1) increases in endotoxin-induced DIC and many thrombotic diseases such as myocardial infarction, type 2 diabetes mellitus, and hyperlipidemia. Anti-PAI-1 monoclonal antibody reduced fibrin deposition in DIC mouse model. Treatment of these monoclonal antibodies for the molecules regulating coagulation-fibrinolysis system may be utilized for acute coronary syndrome and venous thrombosis.     

Helping antibodies. Targeted antithrombotic and fibrinolytic therapy

The development of monoclonal antibodies facilitated an enormous progress in modern medicine in the last years. The targeted inhibition of defined molecular structures allows therapeutic concepts, which before were inconceivable. There are numerous antibodies in clinical use within the area of tumour therapy, chronically inflammatory diseases, transplantation, infections and also in cardiovascular medicine. Different antibody formats are used such as IgG molecules, Fab fragments and single chain antibodies. Single chain antibodies represent the smallest functional form of the antibody and are used preferentially as recombinant antibodies. The therapeutic possibilities of antibody technology are extended by fusion to radioactive or therapeutically active substances. This review focuses on the application of antibodies and fusion proteins as antithrombotic and fibrinolytic drugs. The use of antibodies allows the development of inhibitory agents with clearly defined functional properties, as for example for activation-specific GPIIb/IIIa-blockade. In addition antibodies can be used for targeting antithrombotic and fibrinolytic agents to the thrombus, allowing an effective local action with less bleeding complications.     

Advances in tumour immunotherapy

The clinical goal of tumour immunotherapy is to provide either active or passive immunity against malignancies by harnessing the immune system to target tumours. Although vaccination is an effective strategy to prevent infectious disease, it is less effective in the therapeutic setting for cancer treatment, which might be related to the low immunogenicity of tumour antigens and the reduced immunocompetence of cancer patients. Recent advances in technology have led to the development of passive immunotherapy approaches that utilize the unique specificity of antibodies and T cell receptors to target selected antigens on tumour cells. These approaches are likely to benefit patients and alter the way that clinicians treat malignant disease. In this article we review recent advances in the immunotherapy of cancer, focusing on new strategies to enhance the efficacy of passive immunotherapy with monoclonal antibodies and antigen-specific T cells.     

Gene therapy for malignant liver disease

For most patients with advanced or multifocal hepatocellular carcinoma (HCC) or with metastatic malignant liver disease treatment options are limited, resulting in a poor prognosis. Novel therapeutic strategies such as gene therapy are therefore urgently required. Gene therapeutic approaches use gene delivery systems (vectors) to introduce DNA constructs as therapeutic agents into living cells. Antitumour strategies include the reintroduction of tumour suppressor genes into tumour cells, the expression of foreign enzymes to render tumours susceptible to treatment with chemotherapeutic agents and the enhancement of tumour immunogenicity by expressing immunomodulatory genes or by genetic vaccination with tumour antigens. Furthermore, gene therapy may be also used for anti-angiogenesis to reduce tumour growth and metastatic potential. Other novel approaches aim at the development of genetically altered replication competent viruses, which selectively replicate in tumour cells inducing cell lysis. Although most clinical trials of antitumour gene therapy so far have failed to induce strong therapeutic effects, further improvement of antitumour gene therapy may finally result in potent clinical treatment options for patients with malignant liver tumours.     

Tumor serodiagnosis with monoclonal antibodies

The applicability of monoclonal antibodies (MAb) to the serological diagnosis of solid tumours is discussed. So far test kits with MAb cannot claim higher resolution of specificity as compared with conventional antisera. One approach to solve this problem would be to produce MAb to peptides derived from tumour extracts which are unique to malignant tissue.     

In situ distribution of oncogene products and growth factor receptors in breast carcinoma: c-erbB-2 oncoprotein, EGFr, and PDGFr-beta-subunit

An increasing body of evidence suggests that breast tumour growth is mediated by oncogene products and growth factors which are or which act through cell surface receptors. The aims of the present study were to determine how three of these receptors, c-erbB-2 protein, epidermal growth factor receptor (EGFr) and the beta-subunit of platelet-derived growth factor receptor (PDGFr-beta-subunit), can effectively be demonstrated by immunohistochemical methods in breast tumors, how these receptors are distributed at the cellular level and how their expression correlates with well-established prognostic indicators including hormone receptors and proliferative index. We examined frozen tissue sections of 50 invasive human breast carcinomas, including 45 ductal, four lobular, and one mucinous tumours, by immunocytochemical methods to determine the in situ distributions of c-erbB-2, EGFr, and PDGFr-beta-subunit. We compared staining for c-erbB-2 protein in frozen sections with that in paraffin sections of the same 50 tumours. The immunohistochemical labelling results were compared with tissue hormone receptor content and growth fraction determined by Ki-67 labelling. Strong labelling of tumour cells in frozen sections was detected in 22% of cases, all of the ductal type, stained with rabbit antiserum to c-erbB-2. Labelling for c-erbB-2 protein was generally weaker in paraffin sections than in frozen sections and in six of 11 positive cases, specific staining could be detected only in frozen sections. In immunostains with monoclonal antibody to EGFr, rare cells within tumour were labelled in 60% of the carcinomas. Using a monoclonal antibody to the beta-subunit of PDGFr, consistent labelling of fibrillary cellular processes in the walls of blood vessels and in fibrous stroma around tumour cell nests was detected, but there was no labelling of tumour cells themselves. C-erbB-2 oncoprotein positive tumours were found to be more often oestrogen receptor negative (P less than 0.005) or oestrogen and progesterone receptor negative (P less than 0.01) than c-erbB-2 negative tumours. No significant correlation was observed between c-erbB-2 expression and Ki-67 growth fraction.