Stenting of a renal artery bifurcation stenosis

Gene therapy, in particular the transfer of genes encoding immunostimulatory molecules (cytokines and costimulatory molecules) as well as selectively cytotoxic enzymes and DNA vaccination, has the potential of enhancing cell mediated immune responses against tumours including those of colorectal origin. Genes can be transferred using viral vectors either to cultured tumour cells in vitro that can be returned to the patient as a "cancer vaccine", or directly to tumour cells in vivo. Vaccination with DNA constructs expressing specific tumour antigens characteristic of colorectal neoplasia can trigger immune recognition and destruction of tumour cells. The aim is to tip the balance from protumour to antitumour mechanisms by generating a local immune response and systemic antitumour immune memory to destroy metastases. Studies in murine models, combined with human studies, show that such approaches could become an adjunct to current treatments for human colorectal cancer in the near future.     

GD3/proteosome vaccines induce consistent IgM antibodies against the ganglioside GD3

The gangliosides of melanoma and other tumours of neuroectodermal origin are suitable targets for immune intervention with tumour vaccines. The optimal vaccines in current use contain ganglioside plus bacillus Calmette-Guérin and induce considerable morbidity. We have screened a variety of new adjuvants in the mouse, and describe one antigen-delivery system, proteosomes, which is especially effective. Highly hydrophobic Neisserial outer membrane proteins (OMP) form multimolecular liposome-like vesicular structures termed proteosomes which can readily incorporate amphiphilic molecules such as GD3 ganglioside. The optimal GD3/proteosome vaccine formulation for induction of GD3 antibodies in the mouse is determined. Interestingly, the use of potent immunological adjuvants in addition to proteosomes augments the IgM and IgG antibody titres against OMP in these vaccines but GD3 antibody titres are unaffected. The application of proteosomes to enhance the immune response to GD3 extends the concept of the proteosome immunopotentiating system from lipopeptides to amphipathic carbohydrate epitopes such as cell-surface gangliosides. The demonstrated safety of meningococcal OMP in humans and the data in mice presented here suggest that proteosome vaccines have potential for augmenting the immunogenicity of amphipathic tumour antigens in humans.     

Immunology in the clinical practice. XVI. Paraneoplastic syndromes of the nervous system: pathogenesis and diagnosis

Paraneoplastic neurological syndromes are believed to result from ectopic expression of onconeural antigens by tumours. The resulting immune response is not only directed against the tumour but also cross-reacts with the same or similar antigens in the nervous system. The immune response generates high titred autoantibodies that are associated with specific tumours and neurological syndromes. Paraneoplastic autoantibodies help diagnose neurological syndromes and help direct the search for an underlying tumour. In paraneoplastic syndromes, the course of the tumour is relatively mild. Detection of the autoantibodies might lead to early diagnosis and immunomodulation and anti-tumour treatment before irreversible neuronal cell loss and deficits set in.     

IL-1/IL-3 gene therapy of non-small cell lung cancer (NSCLC) in rats using 'cracked' adenoproducer cells

Cytokine gene therapy was studied in established L42 tumours in syngeneic rats. L42 is a transplantable non-immunogenic non-small cell lung cancer (NSCLC). Genes coding for human interleukin-1 alpha and for rat interleukin-3 beta were transferred by injecting producer cells of recombinant adenovirus vectors into the tumour in attempts to achieve high concentrations of the cytokines inside the tumor without systemic toxicity. Limited tumour growth delay was obtained with viable producer cells. For logistic reasons stocks of pooled frozen producer cells allowed intensive treatment of groups of tumour bearing rats. The cells were lysed by thawing before administration. Ten daily injections of such 'cracked' producer cells induced reproducible tumour responses. These were due to local release of cytokines, not to systemic effects. Growth retardation also occurred in contralateral tumours which were not injected. When rats carrying established tumours were vaccinated with lysates of tumours collected during treatment with 'cracked' producer cells, significant tumour growth retardation was obtained. We speculate that both cytokines, if produced at sufficiently high concentrations in tumours, induce inflammation which in turn initiates an immune response against tumours growing at a distant site. These findings seem to justify further exploration of IL-1 and IL-3 gene transfer for the treatment of cancers.     

Gene-based strategies for the immunotherapy of cancer

T lymphocytes play a crucial role in the host's immune response to cancer. Although there is ample evidence for the presence of tumor-associated antigens on a variety of tumors, they are seemingly unable to elicit an adequate antitumor immune response. Modern cancer immunotherapies are therefore designed to induce or enhance T cell reactivity against tumor antigens. Vaccines consisting of tumor cells transduced with cytokine genes in order to enhance their immunogenicity have been intensely investigated in the past decade and are currently being tested in clinical trials. With the development of novel gene transfer technologies it has now become possible to transfer cytokine genes directly into tumors in vivo. The identification of genes encoding tumor-associated antigens and their peptide products which are recognized by cytotoxic T lymphocytes in the context of major histocompatibility complex class I molecules has allowed development of DNA-based vaccines against defined tumor antigens. Recombinant viral vectors expressing model tumor antigens have shown promising results in experimental models. This has led to clinical trials with replication-defective adenoviruses encoding melanoma-associated antigens for the treatment of patients with melanoma. An attractive alternative concept is the use of plasmid DNA, which can elicit both humoral and cellular immune responses following injection into muscle or skin. New insights into the molecular biology of antigen processing and presentation have revealed the importance of dendritic cells for the induction of primary antigen-specific T cell responses. Considerable clinical interest has arisen to employ dendritic cells as a vehicle to induce tumor antigen-specific immunity. Advances in culture techniques have allowed the generation of large numbers of immunostimulatory dendritic cells in vitro from precursor populations derived from blood or bone marrow. Experimental immunotherapies which now transfer genes encoding tumor-associated antigens or cytokines directly into professional antigen-presenting cells such as dendritic cells are under evaluation in pre-clinical studies at many centers. Gene therapy strategies, such as in vivo cytokine gene transfer directly into tumors as well as the introduction of genes encoding tumor-associated antigens into antigen-presenting cells hold considerable promise for the treatment of patients with cancer.     

Proto-oncogene PML controls genes devoted to MHC class I antigen presentation

Fragments of foreign antigens associated with class I molecules of the major histocompatibility complex (MHC) are presented at the cell surface to elicit an immune response. This presentation requires the coordinated expression of several genes contained in the MHC, including those encoding the MHC class I heavy chain, the proteins LMP-2 and LMP-7, which are involved in the proteasomal degradation of cytosolic antigens into peptide fragments that are destined for association with MHC class I molecules, and TAP-1 and TAP-2, which transport these fragments across the membrane of the endoplasmic reticulum at the start of their journey to the cell surface. In many virus-transformed cell lines and spontaneous tumours, these genes are simultaneously repressed. However, the key factor(s) that are essential for their expression and repression have not been identified. Here we report that the proto-oncogene product PML induces expression of LMP-2, LMP-7, TAP-1 and TAP-2 in an MHC-class I-negative, recurrent tumour, leading to the re-expression of cell-surface MHC in tumours and to rejection of the tumours. PML also regulates MHC expression in untransformed fibroblasts. We conclude that malfunction of PML may enable a tumour to evade the immune defence of its host.     

Induction of antigen presentation by macrophages after phagocytosis of tumour apoptotic cells

Due to their resistance to classical chemotherapies, most human colorectal cancers have a high incidence and a poor prognosis. Immunotherapy using interleukin 2 (IL2) has provided disappointing results in the treatment of these cancers. Recently, however, we have demonstrated that a treatment combining a cell-differentiating agent, sodium butyrate (NaBut) with IL2 resulted in a remission of established peritoneal colorectal carcinomatosis in rats. Separately, neither NaBut nor IL2 treatment cured these tumour-bearing rats. NaBut is known to induce cell differentiation and subsequent apoptosis in epithelial cells, while IL2 stimulates the immune cells capable of participating in tumour rejection. We postulated that the significant therapeutic effect of NaBut/IL2 treatment could be attributed to a NaBut-induced increase in the immunogenicity of the cancer cells. We report here that NaBut induced an apoptotic process in rat colon tumour cells in vivo and in vitro. We observed, in an efficient cure, colocalization of apoptotic bodies and monocytes/macrophages at the periphery of the tumour. We propose that these apoptotic bodies are phagocytosed in vivo by the macrophages. We also showed in vitro that a subpopulation of macrophages involved in the phagocytic clearance of apoptotic cells expresses cell surface molecules associated with antigen presentation and stimulates the proliferation of naive splenocytes. Our data suggest that therapies that recruit massive induction of the apoptotic process in tumour cells could favour tumour antigen presentation via their specific phagocytosis by antigen-presenting cells (APCs). We propose that the development of specific therapies that stimulate both tumour cell apoptosis and the immune system could offer new opportunities in anti-cancer treatments of poorly immunogenic cancer cells.     

Human papillomavirus infection and esophageal cancer: a nationwide seroepidemiologic case-control study in Sweden

BACKGROUND: Despite being immunogenic, gastric cancers overcome antitumour immune responses by mechanisms that have yet to be fully elucidated. Fas ligand (FasL) is a molecule that induces Fas receptor mediated apoptosis of activated immunocytes, thereby mediating normal immune downregulatory roles including immune response termination, tolerance acquisition, and immune privilege. Colon cancer cell lines have previously been shown to express FasL and kill lymphoid cells by Fas mediated apoptosis in vitro. Many diverse tumours have since been found to express FasL suggesting that a "Fas counterattack" against antitumour immune effector cells may contribute to tumour immune escape. AIM: To ascertain if human gastric tumours express FasL in vivo, as a potential mediator of immune escape in stomach cancer. SPECIMENS: Thirty paraffin wax embedded human gastric adenocarcinomas. METHODS: FasL protein was detected in gastric tumours using immunohistochemistry; FasL mRNA was detected in the tumours using in situ hybridisation. Cell death was detected in situ in tumour infiltrating lymphocytes using terminal deoxynucleotidyl transferase mediated dUTP nick end labelling (TUNEL). RESULTS: Prevalent expression of FasL was detected in all 30 resected gastric adenocarcinomas examined. In the tumours, FasL protein and mRNA were co-localised to neoplastic gastric epithelial cells, confirming expression by the tumour cells. FasL expression was independent of tumour stage, suggesting that it may be expressed throughout gastric cancer progression. TUNEL staining disclosed a high level of cell death among lymphocytes infiltrating FasL positive areas of tumour. CONCLUSIONS: Human gastric adenocarcinomas express the immune downregulatory molecule, FasL. The results suggest that FasL is a prevalent mediator of immune privilege in stomach cancer.     

Effect of taurine on angiotensin II-induced hypertrophy of neonatal rat cardiac cells

T Lymphocytes (T cells) recognize short antigenic peptides bound to either MHC I or II molecules, in contrast to antibodies which can bind to native antigen. The mechanism by which antigens are processed into peptides, and the nature of the interactions of antigenic peptides with MHC molecules and with the T cell receptor have now been defined in some detail. Of significance to glycobiologists is the recent appreciation that the carbohydrate of glycoprotein antigens can contribute to the T cell recognition of epitopes presented by MHC molecules. Experiments using model T cell epitopes have demonstrated that carbohydrate can modulate T cell responses in a variety of ways; for example, there are a number of cases where glycopeptide-specific T cell responses have been identified. Many of these glycopeptide-specific T cell responses involve a peptide bearing a single glycosyl residue, thus it appears very likely that both glycan and peptide make contact with the T cell receptor binding site. Significantly, glycopeptide-specific T cell responses have also been detected to native glycoproteins. The ability of carbohydrate to influence T cell recognition of antigen has important consequences for a wide range of immune responses as well as the current strategies for mapping T cell determinants.     

Potentiation of lysis of leukaemia cells by a bispecific antibody to CD33 and CD16 (Fc gamma RIII) expressed by human natural killer (NK) cells

Bispecific antibodies recognizing tumour-associated antigens and trigger molecules expressed on immune effector cells have been shown to redirect cytotoxicity of several types of peripheral blood cells against relevant tumour targets. Among various effector cells, natural killer (NK) cells appear to play a role in defence against leukaemia. Here we report the successful chemical conjugation of monoclonal antibodies to CD33 and CD16 to create a bispecific antibody (BsAb 251 x 3G8). This bispecific antibody is capable of augmenting the killing of otherwise resistant leukaemia cells by peripheral blood lymphocytes (PBL), purified resting NK (R-NK) cells, and activated NK (A-NK) cells. BsAb 251 x 3G8 may play a role in the therapy of acute myeloid leukaemia (AML) through redirecting the cytotoxic activity of endogenous or adoptively transferred NK cells.     

Cellular vaccines

This project is devoted to the development of novel cellular vaccines designed to treat cancer patients. These cellular vaccines present and enhance immunogens, which will elicit a potent immune response. The goal is to achieve safe and effective immune reaction against the patient's own tumour. (1) Autologous cellular vaccines are prepared by processing circulating blood mononuclear cells outside of the patient's body (ex vivo) to differentiate them into antigen-presenting cells (APCs). Monocyte-derived APCs (MD-APCs) are then grown in the presence of exogenous target antigens (tumour cell debris, or apoptotic bodies) to become fully mature APCs. (2) Functionality for antigen presentation to T cells of ex vivo MD-APCs is evaluated in vivo. (3) Cellular vaccines are tested in selected rodent animal models. Efficiency and immune response are monitored in pertinent experimental systems for cancer. Pharmacological data are generated for clinical investigation. Tolerance and biologic effects are documented in primates. (4) The first clinical trials on cancer patients are taking place in 1998 on melanoma and prostate cancer to validate the concept. Specialized cell processors with dedicated software and standardized controls are being developed and used for the preparation of cellular vaccines. (5) The evaluation of new non-viral vectors and the validation of new non-viral transfection methods of mononuclear cells with marker genes is in progress and will lead to the ex vivo transfection of genes coding for immunostimulating cytokines or for tumour antigens in MD-APCs. Efficiency will be validated in vitro and in animal models. The ex vivo and animal model studies validate the clinical relevance of this new cellular immunotechnology. Clinical validation of individual autologous cellular vaccines in specific indications for which no treatment is presently available will allow the development of cellular and gene immunotherapy for other types of cancers.     

Epidermal growth factor receptor and its inhibition in radiotherapy: in vivo findings

Increasing evidence shows that dysregulated epidermal growth factor receptor (EGFR) signalling plays an important part in neoplasia. When over expressed or mutated, EGFR is frequently associated with more aggressive tumour growth, poor patient prognosis and resistance of tumours to cytotoxic agents, including radiation. The present studies with murine carcinomas showed that there is an inverse correlation between the level of EGFR and tumour radiocurability. Likewise, the present clinical study in patients with head and neck cancer shows that EGFR over expression correlates with poorer tumour response to radiotherapy. Adding EGFR to tumour cells in vitro protected cells against the cytotoxic action of radiation, whereas blocking EGFR with anti-EGFR antibodies enhanced cell radiosensitivity. A casual relationship between EGFR and increased cellular resistance to radiation was established by transferring the EGFR gene into low EGFR-expressing radiosensitive tumour cells, which then become radioresistant. Radiation activated EGFR and its downstream signalling pathways in radioresistant but not in radiosensitive tumours, and this effect was associated with increased resistance to radiation, and enhanced repopulation in irradiated tumours. Increasing evidence shows that blockage of EGFR or interference with any of the steps in its signal transduction cascade can counteract negative outcomes of EGFR signalling, which has recently been explored as a therapeutic strategy in cancer treatment. The present findings demonstrate that treatment of human tumour xenografts with C225, an anti-EGFR monoclonal antibody, dramatically enhanced tumour response to radiation. Overall, the findings show that over expression of EGFR may serve as a predictor of tumour treatment outcome by radiotherapy and as a therapeutic target to enhance the efficacy of radiotherapy.     

Antigen processing/presenting and oncogenesis

It has been established in recent years that a number of tumor cell types express tumor antigens, yet the host's immune system fails to recognize them. The antigen processing/presenting machinery, which plays a crucial role in generating an immune response, and possible causes for its inability of processing/presenting tumor antigens are reviewed. These causes are related to the expression of major histocompatibility complex molecules, costimulatory molecules, and tumor antigens by tumor cells, and the types of cytokines produced. Therapeutic measures include transfecting tumor cells with genes that encode major histocompatibility complex and costimulatory molecules, cytokines, and tumor antigens. In addition, tumor peptide vaccines are evaluated. However, developing an immune response to tumor antigens carries with it the risk of autoimmune disease.     

Tumor-reactive immunoglobulins in ovarian cancer: diagnostic and therapeutic significance? (review)

Inhibition of the immune system has been observed in association with most stages of ovarian cancer; however, the mechanisms involved in the induction and maintenance of this chronic immune unresponsiveness associated with cancer progression are poorly understood. This immunosuppressed state is primarily defined as the failure to eradicate the tumor. This immunosuppressed state is generally associated with decreased numbers and reactivity of lymphoid cells in women with ovarian cancer. The degree of immune dysfunction in ovarian cancer patients has been demonstrated to correlate with patient survival. While ovarian cancer patients generally fail to exhibit effective immunosurveillance, as manifested by continued tumor growth and progression, the presence of tumor-reactive immunoglobulins can be demonstrated in these women, indicating the continued presence of immune recognition. We have not only demonstrated the presence of tumor-reactive antibodies in ovarian cancer patients, but have also shown that the levels of these antibodies increase as the disease progresses. The antigens recognized by the patients' humoral response have been identified as either membrane-associated or intra-cellular. In general, the localization of these antigens tend to be linked to the patient's prognosis. The presence of a humoral response against intracellular proteins are correlated with poor prognosis, while autoantibodies reactive with surface components appear to have a better prognosis. In addition to general antigen recognition, these reactive antibodies have been utilized to define specific epitopes on tumor-associated proteins. Certain specific antigenic epitopes exhibit common recognition among patients with the same tumor type. The specific recognition of certain epitopes can provide early evidence of aberrant protein expression and this aberrant expression of certain proteins, such as procathepsin D, appear to be linked to the tumor's acquisition of specific malignant characteristics, including metastasis formation and chemoresistance. Despite the existence of circulating tumor-reactive immunoglobulins, their presence correlates, in general, with poor prognosis and poor host survival. Since tumor-reactive immunoglobulins are elicited and can be detected early in the development of tumors and their enhanced synthesis is induced prior to the clinical manifestation of recurrence, the assessment of the tumor-reactive immune response against specific antigenic epitopes should represent an early significant diagnostic and prognostic marker in ovarian cancer.     

Breast cancer immunotherapy: current status and future prospects

The development of an immunotherapeutic approach to cancer is the concern for many immunologists, but despite the impressive progress over the past decade, such as the identification of tumour antigens and antigenic peptides as potential targets, there are still many obstacles in eliciting an effective immune response to eradicate cancer. Mucins have attracted interest as potential targets for immunotherapy in the development of vaccines for cancers expressing Mucin1 (MUC1; e.g. breast, pancreas, ovary etc.). All of the identified targets for cancer, including MUC1, are normal proteins; however MUC1 expressed on tumours can be considered as tumour specific due to their overexpression, altered glycosylation and its ubiquitous distribution on the cell surface rather than at the secretory pole in adenocarcinomas. These observations have led to the development of several different approaches to immunize against breast cancer using synthetic carbohydrates or peptides conjugated to carriers and given together with a variety of adjuvants to elicit the appropriate immune response. Mannan, a polymannose carbohydrate isolated from the cell wall of yeast, is an appropriate and effective protein carrier for eliciting a cellular (T1-type) or humoral (T2-type) immune response depending on the mode of conjugation (oxidized or reduced). In addition, mannan holds promise and opens many avenues as a carrier for vaccine development for other antigens. Several clinical trials are in progress to evaluate the immunogenicity of MUC1 and its suitability as to use for immunotherapy/vaccine for breast cancer.     

Role of the vascular component in growth of solid tumors: a historical review

Angiogenesis is a fundamental biological process by which new capillary blood vessels are formed. It is essential in many physiological conditions, such as embryonic development, ovulation and wound repair, and pathological ones, such as arthritis, diabetic retinopathy, and tumours. Solid tumours have angiogenesis capacity, and tumour growth and metastasis are angiogenesis-dependent. Neoplastic cell populations can grow to form a clinically evident tumour only if the host produces a vascular network sufficient to sustain tumour growth. Furthermore, the new blood vessels provide a gateway for tumour cells to enter the circulation and metastasize to distant sites. Tumour angiogenesis is essentially mediated by angiogenic molecules elaborated by tumour cells. We review here the most important literature on this topic and emphasize the crucial and paradigmatic role of this biological process and its relevance in a possible anti-angiogenic therapeutic approach to the treatment of solid tumours.     

Endothelial-cell proliferation in experimental tumours

The proliferation characteristics of vascular endothelium have been studied in 131 individual experimental tumours, representing 18 transplanted tumour lines. The labelling index (LI) is high in most tumours, with a mean value of 0.9%, regardless of the growth rate of the tumours, or whether different tumour types are considered or individual tumours from within one line are studied in detail. A similar high LI value has been found by others for a human tumour. These high LI values may even underestimate the proliferation in new capillary buds. The high proliferative index of tumour endothelium is in marked contrast with the previously reported low 3HTdR uptake into normal tissue blood vessels. It seems likely that it is the type of new vessels formed that will influence tumour growth rates more than the simple rate of endothelial-cell proliferation. The large difference between the proliferation characteristics of tumour endothelium and normal tissue endothelium, recently identified as a possible approach for tumour therapy, has now been confirmed for a range of animal tumours and a human tumour.     

PRAME, a gene encoding an antigen recognized on a human melanoma by cytolytic T cells, is expressed in acute leukaemia cells

Gene PRAME was found to encode an antigen recognized on a human melanoma cell line by an autologous cytolytic T-lymphocyte clone. This gene is expressed at a high level in a very large fraction of tumours, such as melanomas, non-small-cell lung carcinomas, sarcomas, head and neck tumours and renal carcinomas. It is therefore a candidate for tumour immunotherapy even though some low expression is found in certain normal tissues. We tested by RT-PCR the expression of PRAME on more than 250 bone marrow or blood samples from patients with a haematological malignancy. Approximately 25% of the acute leukaemia samples were positive. Remarkably, all acute myeloblastic leukaemias that carried the chromosomal translocation t(8;21), which fuses the genes AML1 and ETO, expressed PRAME at a high level.     

Phenotypic analysis of resident lymphoid cells in the conjunctiva and adnexal tissues of rat

The conjunctival associated lymphoid tissue is considered to be an integral part of the mucosal immune system. Under normal circumstances immune mechanisms in mucosal associated lymphoid tissue of the gut and bronchus can selectively suppress, rather than enhance, immune responsiveness to encountered antigens, inducing a state of tolerance. It is possible that conjunctival associated lymphoid tissue can also induce a state of tolerance to encountered antigens. Such a response may be exploited to modulate immune mediated ocular disease. Enhanced tolerance may protect the host against foreign antigen. Alternatively, under certain circumstances when the normal immune system is altered or disrupted the mucosal tissue may act to induce sensitisation and trigger immune mediated disease. The rat is frequently used as an animal model of immune mediated eye disease, but the normal profile of immune cells in the rat conjunctiva has not been studied. This information is essential for meaningful interpretation in the experimental situation. In this study we examined the immunophenotype of lymphoid tissue associated with the conjunctiva, lacrimal gland and Harderian gland of the Lewis rat. CD4+, Ia+ and the monocyte/macrophage population of cells were found predominantly in the substantia propria of the conjuctiva and interstitial connective tissue of the glands. CD8+ cells were distributed mainly in relation to the conjunctival and glandular epithelium. Goblet cells stained strongly with the monoclonal antibody (MAb) MRC OX-39, which is a marker for IL-2 receptors. The overall pattern of distribution of immunocompetent cells in the rat was found to be similar to that reported in humans.     

SWEEPS: a program for the acquisition and analysis of neurophysiological data

The metastatic spread of solid tumours is responsible directly or indirectly for most cancer-related deaths. Our understanding of the molecular genetic and biological events that contribute to tumor cell dissemination has increased considerably over the last decade. It is clear that close anatomic and temporal co-operation between cellular adhesion molecules, extracellular matrix (ECM)-degrading proteases and peptides inducing tumour vascularisation are essential components of the metastatic behaviour of cancer cells. Although this enhanced understanding may have little immediate impact on patient survival (about 50% of patients have established metastatic disease at first presentation), it has led to the development of novel anti-metastatic therapies targeting distinct molecules and steps in the metastatic cascade. Here we review the role of matrix-degrading enzymes, changes in cellular adhesive capacity and tumour angiogenesis during cancer spread, highlighting areas that are of emerging importance in the clinic.