cefoxitin and Neoplasms

While the underlying causes of cancer are genetic modifications, changes in cellular states mediate cancer development. Tumor cells display markedly changed glycosylation states, of which the O-GalNAc glycans called the Tn and TF antigens are particularly common. How these antigens get over-expressed is not clear. The expression levels of glycosylation enzymes fail to explain it.. We describe the regulation of O-GalNAc glycosylation initiation and extension with emphasis on the initiating enzymes ppGalNAcTs (GALNTs), and introduce the GALA pathway--a change in GALNTs compartmentation within the secretory pathway that regulates Tn levels. We discuss the roles of O-GalNAc glycans and GALNTs in tumorigenic processes and finally consider diagnostic and therapeutic perspectives.. Contrary to a common hypothesis, short O-glycans in tumors are not the result of an incomplete glycosylation process but rather reveal the activation of regulatory pathways. Surprisingly, high Tn levels reveal a major shift in the O-glycoproteome rather than a shortening of O-glycans. These changes are driven by membrane trafficking events.. Many attempts to use O-glycans for biomarker, antibody and therapeutic vaccine development have been made, but suffer limitations including poor sensitivity and/or specificity that may in part derive from lack of a mechanistic understanding. Deciphering how short O-GalNAc glycans are regulated would open new perspectives to exploit this biology for therapeutic usage. This article is part of a Special Issue entitled "Glycans in personalised medicine" Guest Editor: Professor Gordan Lauc.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Cancer Vaccines; Galactosamine; Glycoproteins; Glycosylation; Humans; Neoplasm Proteins; Neoplasms; Oligosaccharides

Glycosylation is one of the major posttranslational modifications of proteins. N-glycosylation (Asn-linked) and O-glycosylation (Ser/Thr-linked) are the two main forms. Abnormal O-glycosylation is frequently observed on the surface of tumor cells, and is associated with an adverse outcome and poor prognosis in patients with cancer. O-glycans (Tn, sTn, and T antigen) can be synthesized in the Golgi apparatus with the aid of several glycosyltransferases (such as T-synthase and ST6GalNAc-I) in a suitable environment. The unique molecular chaperone of T-synthase is Cosmc, which helps T-synthase to fold correctly in the endoplasmic reticulum. Dysregulation of these glycosyltransferases, molecular chaperones, or the environment is involved in the dysregulation of O-glycans. Tn, sTn, and T antigen neo- or over-expression occurs in many types of cancer including gastric, colon, breast, lung, esophageal, prostate, and endometrial cancer. This review discusses the major synthetic pathway of O-glycans and the mechanism by which Tn, sTn, and T antigens promote tumor metastasis.

Topics: Antigens, Tumor-Associated, Carbohydrate; Antigens, Viral, Tumor; Carbohydrate Sequence; Endoplasmic Reticulum; Galactosyltransferases; Gene Expression Regulation, Neoplastic; Glycosylation; Golgi Apparatus; Humans; Molecular Chaperones; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Protein Folding; Protein Processing, Post-Translational; Sialyltransferases; Tumor Cells, Cultured

The carbohydrate antigens Tn and sialyl-Tn (STn) are expressed in most carcinomas and usually absent in healthy tissues. These antigens have been correlated with cancer progression and poor prognosis, and associated with immunosuppressive microenvironment. Presently they are used in clinical trials as therapeutic vaccination, but with limited success due to their low immunogenicity. Alternatively, anti-Tn and/or STn antibodies may be used to harness the immune system against tumor cells. Whilst the development of antibodies against these antigens had a boost two decades ago for diagnostic use, so far no such antibody entered into clinical trials. Possible limitations are the low specificity and efficiency of existing antibodies and that novel antibodies are still necessary. The vast array of methodologies available today will allow rapid antibody development and novel formats. Following the advent of hybridoma technology, the immortalization of human B cells became a methodology to obtain human monoclonal antibodies with better specificity. Advances in molecular biology including phage display technology for high throughput screening, transgenic mice and more recently molecularly engineered antibodies enhanced the field of antibody production. The development of novel antibodies against Tn and STn taking advantage of innovative technologies and engineering techniques may result in innovative therapeutic antibodies for cancer treatment.

Topics: Animals; Antibodies; Antigens, Tumor-Associated, Carbohydrate; Genetic Engineering; Humans; Immunotherapy; Neoplasms

C-type lectin receptors (CLRs) on antigen-presenting cells (APCs) facilitate uptake of carbohydrate antigens for antigen presentation, modulating the immune response in infection, homeostasis, autoimmunity, allergy, and cancer. In this review, we focus on the role of the macrophage galactose type C-type lectin (MGL) in the immune response against self-antigens, pathogens, and tumor associated antigens (TAA). MGL is a CLR exclusively expressed by dendritic cells (DCs) and activated macrophages (MØs), able to recognize terminal GalNAc residues, including the sialylated and nonsialylated Tn antigens. We discuss the effects on DC function induced throughout the engagement of MGL, highlighting the importance of the antigen structure in the modulation of immune response. Indeed modifying Tn-density, the length, and steric structure of the Tn-antigens can result in generating immunogens that can efficiently bind to MGL, strongly activate DCs, mimic the effects of a danger signal, and achieve an efficient presentation in HLA classes I and II compartments.

Topics: Acetylgalactosamine; Antigen Presentation; Antigens, Bacterial; Antigens, Tumor-Associated, Carbohydrate; Antigens, Viral; Autoantigens; Dendritic Cells; Gene Expression; HLA Antigens; Humans; Immunologic Factors; Lectins, C-Type; Ligands; Macrophages; Neoplasms; Peptidomimetics; Signal Transduction

The Tn antigen is a tumor-associated carbohydrate antigen that is not normally expressed in peripheral tissues or blood cells. Expression of this antigen, which is found in a majority of human carcinomas of all types, arises from a blockage in the normal O-glycosylation pathway in which glycans are extended from the common precursor GalNAcα1-O-Ser/Thr (Tn antigen). This precursor is generated in the Golgi apparatus on newly synthesized glycoproteins by a family of polypeptide α-N-acetylgalactosaminyltransferases (ppGalNAcTs) and then extended to the common core 1 O-glycan Galβ1-3GalNAcα1-O-Ser/Thr (T antigen) by a single enzyme termed the T-synthase (core 1 β3-galactosyltransferase or C1GalT). Formation of the active form of the T-synthase requires a unique molecular chaperone termed Cosmc, encoded by Cosmc on the X-chromosome (Xq24 in humans, Xc3 in mice). Cosmc resides in the endoplasmic reticulum (ER) and prevents misfolding, aggregation, and proteasome-dependent degradation of newly synthesized T-synthase. Loss of expression of active T-synthase or Cosmc can lead to expression of the Tn antigen, along with its sialylated version Sialyl Tn antigen as observed in several cancers. Both genetic and epigenetic pathways, in addition to potential metabolic regulation, can result in abnormal expression of the Tn antigen. Engineered expression of the Tn antigen by disruption of either C1GalT (T-syn) or Cosmc in mice is associated with a tremendous range of pathologies and engineered expression of the Tn antigen in mouse embryos leads to embryonic death. Studies indicate that many membrane glycoproteins expressing the Tn antigen and/or truncated O-glycans may be dysfunctional, due to degradation and/or misfolding. Thus, expression of normal O-glycans is associated with health and homeostasis whereas truncation of O-glycans, e.g. the Tn and/or Sialyl Tn antigens is associated with cancer and other pathologies.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Carbohydrate Sequence; Glycosylation; Humans; Mice; Neoplasms; Polysaccharides

Tumor cells exhibit striking changes in cell surface glycosylation as a consequence of dysregulated glycosyltransferases and glycosidases. In particular, an increase in the expression of certain sialylated glycans is a prominent feature of many transformed cells. Altered sialylation has long been associated with metastatic cell behaviors including invasion and enhanced cell survival; however, there is limited information regarding the molecular details of how distinct sialylated structures or sialylated carrier proteins regulate cell signaling to control responses such as adhesion/migration or resistance to specific apoptotic pathways. The goal of this review is to highlight selected examples of sialylated glycans for which there is some knowledge of molecular mechanisms linking aberrant sialylation to critical processes involved in metastasis.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Cell Movement; Glycosylation; Humans; Integrins; Lewis X Antigen; N-Acetylneuraminic Acid; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasms; Phenotype; Polysaccharides; Sialyl Lewis X Antigen

Glycoproteins in animal cells contain a variety of glycan structures that are added co- and/or posttranslationally to proteins. Of over 20 different types of sugar-amino acid linkages known, the two major types are N-glycans (Asn-linked) and O-glycans (Ser/Thr-linked). An abnormal mucin-type O-glycan whose expression is associated with cancer and several human disorders is the Tn antigen. It has a relatively simple structure composed of N-acetyl-D-galactosamine with a glycosidic α linkage to serine/threonine residues in glycoproteins (GalNAcα1-O-Ser/Thr), and was one of the first glycoconjugates to be chemically synthesized. The Tn antigen is normally modified by a specific galactosyltransferase (T-synthase) in the Golgi apparatus of cells. Expression of active T-synthase is uniquely dependent on the molecular chaperone Cosmc, which is encoded by a gene on the X chromosome. Expression of the Tn antigen can arise as a consequence of mutations in the genes for T-synthase or Cosmc, or genes affecting other steps of O-glycosylation pathways. Because of the association of the Tn antigen with disease, there is much interest in the development of Tn-based vaccines and other therapeutic approaches based on Tn expression.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Galactosyltransferases; Glomerulonephritis, IGA; Humans; Mice; Molecular Chaperones; Neoplasms; Polysaccharides

Many evolving treatments for cancer patients are based on the targeted delivery of therapeutic cargoes to and into cancer cells. The advent of monoclonal antibodies and the use of peptide hormones, growth factors and cytokines have historically provided a spectrum of ligands needed to selectively target tumor-associated antigens on cancer cells. However, issues linked to the size, cost and immunogenicity of protein-based ligands have led to the search for alternate ligand families. The advent of short synthetic oligonucleotide ligands known as aptamers now provides a simple strategy to select for membrane-impermeant aptamers tailored to precisely target internalized surface markers present on cancer cells. Here we described how 25-base long, synthetic single-stranded DNA aptamers were derived to bind to known internalized tumor markers such as CD33, CEA, MUC1 and Tn antigens and are imported through these surface portals into cancer cells. The key consequence of using internalized aptamers is their ability to accumulate inside the cells, thus routing their therapeutic cargoes to intracellular sites relevant to their action. Internalized aptamers are discussed in the context of how such ligands have been used to create a range of guided therapeutic agents ranging from drug-based conjugates up to targeted nanoparticles.

Topics: Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Antigens, Tumor-Associated, Carbohydrate; Aptamers, Nucleotide; Cell Membrane; Drug Delivery Systems; Humans; Mucin-1; Neoplasm Proteins; Neoplasms; Protein Binding; Receptors, Cell Surface; Sialic Acid Binding Ig-like Lectin 3

Topics: Antibody Formation; Antigens, Tumor-Associated, Carbohydrate; Antigens, Viral, Tumor; Immunity, Cellular; Neoplasms; Prognosis

Aberrant glycosylation is a hallmark of cancer cells. The blood group precursors T (Thomsen-Friedenreich) and Tn epitopes are shielded in healthy and benign-diseased tissues but uncovered in approx. 90% of carcinomas. T and Tn glycoproteins are specific, autoimmunogenic pancarcinoma antigens. These antigens may also be found in neoplastic blood cells (and on LTV-II infected T lymphocytes). Fundamental chemical and physical aspects of these glycoproteins of primary carcinomas are discussed first. Tn and T epitopes are cell and tissue adhesion molecules, essential in invasion by and metastasis of carcinoma which includes adherent and proliferative phases. These properties are then delineated next, followed by consideration of pathophysiological and clinical aspects of these antigens. Immunohistochemical studies of the extent of Tn antigen expression in primary breast carcinoma demonstrate it highly significant correlation with clinicopathological tumor stages, and hence its value as a reliable prognosticator. On the other hand, there is no significant, prognostically useful association between T antigen expression and clinical disease course. Everyone has "preexisting" anticarcinoma anti-Tn and anti-T antibodies, induced predominantly by the intestinal flora, while cellular immune responses to T and Tn epitopes are evoked only by carcinomas and some lymphomas. Carcinoma dedifferentiation leading to predominance of Tn over T epitopes is described, as is the role of Tn and T epitopes in very early, including preclinical, carcinoma detection. The highest sensitivities in carcinoma detection are for preclinical and the earliest clinical stages. Obviously, preclinical carcinoma detection is of practical importance. T/anti-T tests detected preclinical carcinoma in 77% of 48 patients long (mean 6 years) before their biopsy/X-ray results became positive. There were no false predictions of carcinoma in 38 control persons with benign diseases (observation average 4.8 years). These findings open a novel window for both curative approaches and pathophysiological studies. The autoimmunogenicity of carcinoma T/Tn antigen led us more than two decades ago to begin intradermal vaccination of patients with advanced breast carcinoma of stages IV-IIb, predominately after modified radical mastectomy and sometimes lumpectomy plus axillary dissection always followed by adjuvant radio/chemotherapy. The vaccine consists of human group O red blood cell membrane derived, HLA-free T/T

Topics: Antigens, Tumor-Associated, Carbohydrate; Epitopes; Humans; Neoplasm Invasiveness; Neoplasm Metastasis; Neoplasms; Prognosis

Tn and sialyl-Tn carbohydrate structures, first identified in glycophorins of persons with the rare Tn syndrome, were found to be present on the surface of most cancer cells. In this article, the studies on Tn and sialyl-Tn antigens as diagnostic and prognostic tumor markers and as immunogens in vaccines for cancer immunotherapy are shortly reviewed.

Topics: Antibodies, Monoclonal; Antigens, Tumor-Associated, Carbohydrate; Biomarkers, Tumor; Female; Humans; Male; Neoplasms; Prognosis; Vaccination

Topics: Animals; Antibodies, Monoclonal; Antibodies, Neoplasm; Antibody Specificity; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Autoantibodies; Autoantigens; Biomarkers, Tumor; Carbohydrate Sequence; Disaccharides; Epitopes; Evaluation Studies as Topic; Humans; Immunologic Tests; Lectins; Molecular Sequence Data; Neoplasm Invasiveness; Neoplasms; Neoplasms, Experimental; Organ Specificity; Prognosis

Topics: Antibodies, Monoclonal; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Biomarkers, Tumor; Carbohydrate Sequence; Disaccharides; Glycolipids; Humans; Lewis X Antigen; Molecular Sequence Data; Neoplasms

The results of three consecutive clinical trials on the therapy of anaerobic infections in cancer patients have been compared. The success rate with lamoxactam (94%) (6000mg/d i.v.) was statistically different from that of doxycycline (63%) (300 mg/d per os). Clindamycin (1200 mg/d per os), clindamycin (2700 mg/d i.v.) and cefoxitin (6000 mg/d i.v.) resulted in a favourable outcome in approximately 80% of the patients. Mixed anaerobic infections had a similar response rate (80%). Even when the anaerobic pathogen was resistant to therapy, six of ten patients were cured. Surgical drainage played an important role, but was difficult to assess precisely.

Topics: Anti-Bacterial Agents; Bacterial Infections; Cefoxitin; Cephamycins; Clindamycin; Clinical Trials as Topic; Doxycycline; Drainage; Female; Humans; Male; Middle Aged; Moxalactam; Neoplasms; Retrospective Studies; Tinidazole

Clindamycin and cefoxitin with or without gentamicin were administered to cancer patients having localized infections presumably caused by anaerobic pathogens. The rates of favorable response were 89% in patients receiving clindamycine alone and 78% in patients receiving cefoxitin alone. When the total experience is considered (clindamycin or cefoxitin with and without gentamicin), 20 of 24 patients (83%) responded to clindamycin and 18 of 22 (82%) responded to cefoxitin. Both therapies were well tolerated. Clindamycin was found to be more effective than cefoxitin in eradicating the offending anaerobic pathogens from the site of infection. Aerobic pathogens were frequently isolated along with anaerobes from the infectious sites in this series; their susceptibility or resistance to clindamycin or cefoxitin did not influence the therapeutic response.

Topics: Anaerobiosis; Bacterial Infections; Cefoxitin; Clindamycin; Female; Humans; Male; Middle Aged; Neoplasms

Glycoproteins with diverse glycans are essential to human cells, and subtle differences in glycan structures may result in entirely different functions. One typical example is proteins modified with O-linked β-

Topics: Acetylgalactosamine; Antigens, Tumor-Associated, Carbohydrate; Glycoproteins; Humans; Mass Spectrometry; Neoplasms

Improving outcomes for cancer patients during treatment and monitoring for cancer recurrence requires personalized care which can only be achieved through regular surveillance for biomarkers. Unfortunately, routine detection for blood-based biomarkers is cost-prohibitive using currently specialized laboratories. Using a rapid self-assembly sensing interface amenable to methods of mass production, we demonstrate the ability to detect and quantify a small carbohydrate-based cancer biomarker, Tn antigen (αGalNAc-Ser/Thr) in a small volume of blood, using a test format strip reminiscent of a blood glucose test. The detection of Tn antigen at picomolar levels is achieved through a new transduction mechanism based on the impact of Tn antigen interactions on the molecular dynamic motion of a lectin cross-linked lubricin antifouling brush. In tests performed on retrospective blood plasma samples from patients presenting three different tumor types, differentiation between healthy and diseased patients was achieved, highlighting the clinical potential for cancer monitoring.

Topics: Carbohydrates; Humans; Neoplasms; Point-of-Care Systems; Retrospective Studies

Novel immunotherapies targeting cancer-associated truncated O-glycans Tn (GalNAcα-Ser/Thr) and STn (Neu5Acα2-6GalNacα-Ser/Thr) are promising strategies for cancer treatment. However, no comprehensive, antibody-based mapping of truncated O-glycans in tumours exist to guide drug development.. We used monoclonal antibodies to map the expression of truncated O-glycans in >700 tissue cores representing healthy and tumour tissues originating from breast, colon, lung, pancreas, skin, CNS and mesenchymal tissue. Patient-derived xenografts were used to evaluate Tn expression upon tumour engraftment.. The Tn-antigen was highly expressed in breast (57%, n = 64), colorectal (51%, n = 140) and pancreatic (53%, n = 108) tumours, while STn was mainly observed in colorectal (80%, n = 140) and pancreatic (56%, n = 108) tumours. We observed no truncated O-glycans in mesenchymal tumours (n = 32) and low expression of Tn (5%, n = 87) and STn (1%, n = 75) in CNS tumours. No Tn-antigen was found in normal tissue (n = 124) while STn was occasionally observed in healthy gastrointestinal tissue. Surface expression of Tn-antigen was identified across several cancers. Tn and STn expression decreased with tumour grade, but not with cancer stage. Numerous xenografts maintained Tn expression.. Surface expression of truncated O-glycans is limited to cancers of epithelial origin, making Tn and STn attractive immunological targets in the treatment of human carcinomas.

Topics: Animals; Antibodies, Monoclonal; Antigens, Tumor-Associated, Carbohydrate; Case-Control Studies; Down-Regulation; Gene Expression Regulation, Neoplastic; Humans; Mice; Neoplasm Grading; Neoplasm Staging; Neoplasm Transplantation; Neoplasms; Tissue Array Analysis; Up-Regulation

Core 1-derived mucin-type O-glycans (O-glycans) are a major component of gastric mucus with an unclear role. To address this, we generated mice lacking gastric epithelial O-glycans (GEC C1galt1-/-). GEC C1galt1-/- mice exhibited spontaneous gastritis that progressed to adenocarcinoma with ∼80% penetrance by 1 yr. GEC C1galt1-/- gastric epithelium exhibited defective expression of a major mucus forming O-glycoprotein Muc5AC relative to WT controls, which was associated with impaired gastric acid homeostasis. Inflammation and tumorigenesis in GEC C1galt1-/- stomach were concurrent with activation of caspases 1 and 11 (Casp1/11)-dependent inflammasome. GEC C1galt1-/- mice genetically lacking Casp1/11 had reduced gastritis and gastric cancer progression. Notably, expression of Tn antigen, a truncated form of O-glycan, and CASP1 activation was associated with tumor progression in gastric cancer patients. These results reveal a critical role of O-glycosylation in gastric homeostasis and the protection of the gastric mucosa from Casp1-mediated gastric inflammation and cancer.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Carcinogenesis; Caspase 1; Female; Gastric Mucosa; Gastritis; Glycosylation; Homeostasis; Humans; Inflammation; Male; Mice; Mucins; Mucus; Neoplasms; Polysaccharides; Stomach Neoplasms

To understand the role of human natural IgM known as antibodies against the carbohydrate epitope Tn, the antibodies were isolated using GalNAcα-Sepharose affinity chromatography, and their specificity was profiled using microarrays (a glycan array printed with oligosaccharides and bacterial polysaccharides, as well as a glycopeptide array), flow cytometry, and inhibition ELISA. The antibodies bound a restricted number of GalNAcα-terminated oligosaccharides better than the parent monosaccharide, e.g., 6-O-Su-GalNAcα and GalNAcα1-3Galβ1-3(4)GlcNAcβ. The binding with several bacterial polysaccharides that have no structural resemblance to the affinity ligand GalNAcα was quite unexpected. Given that GalNAcα is considered the key fragment of the Tn antigen, it is surprising that these antibodies bind weakly GalNAcα-OSer and do not bind a wide variety of GalNAcα-OSer/Thr-containing mucin glycopeptides. At the same time, we have observed specific binding to cells having Tn-positive glycoproteins containing similar glycopeptide motifs in a conformationally rigid macromolecule. Thus, specific recognition of the Tn antigen apparently requires that the naturally occurring "anti-Tn" IgM recognize a complex epitope comprising the GalNAcα as an essential component and a fairly long amino acid sequence where the amino acids adjacent to GalNAcα do not contact the antibody paratope; i.e., the antibodies recognize a spatial epitope or a molecular pattern rather than a classical continuous sequence. In addition, we have not found any increase in the binding of natural antibodies when GalNAcα residues were clustered. These results may help in further development of anticancer vaccines based on synthetic Tn constructs.

Topics: Amino Acid Sequence; Antibody Affinity; Antibody Specificity; Antigen-Antibody Reactions; Antigens, Tumor-Associated, Carbohydrate; Carbohydrate Sequence; Epitopes; Humans; Immunity, Innate; Immunoglobulin M; Jurkat Cells; Neoplasms

Granulysin is a protein present in the granules of human cytotoxic T lymphocytes (CTL) and natural killer (NK) cells, with cytolytic activity against microbes and tumors. Previous work demonstrated the therapeutic effect of the intratumoral injection of recombinant granulysin and of the systemic injection of an immunotoxin between granulysin and the anti-carcinoembryonic antigen single-chain Fv antibody fragment MFE23, which were produced in the yeast

Topics: A549 Cells; Antigens, Differentiation, T-Lymphocyte; Antigens, Tumor-Associated, Carbohydrate; Cell Line, Tumor; Cell Proliferation; Cell Survival; Electroporation; Humans; Immunotoxins; Jurkat Cells; MCF-7 Cells; Neoplasms; Protein Engineering; Recombinant Proteins; Saccharomycetales; Single-Chain Antibodies

Human cancer cells were eradicated by adoptive transfer of T cells transduced with a chimeric antigen receptor (CAR) made from an antibody (237Ab) that is highly specific for the murine Tn-glycosylated podoplanin (Tn-PDPN). The objectives were to determine the specificity of these CAR-transduced T (CART) cells and the mechanism for the absence of relapse. We show that although the 237Ab bound only to cell lines expressing murine Tn-PDPN, the 237Ab-derived 237CART cells lysed multiple different human and murine cancers not predicted by the 237Ab binding. Nevertheless, the 237CART cell reactivities remained cancer specific because all recognitions were dependent on the Tn glycosylation that resulted from COSMC mutations that were not present in normal tissues. While Tn was required for the recognition by 237CART, Tn alone was not sufficient for 237CART cell activation. Activation of 237CART cells required peptide backbone recognition but tolerated substitutions of up to 5 of the 7 amino acid residues in the motif recognized by 237Ab. Together, these findings demonstrate what we believe is a new principle whereby simultaneous recognition of multiple independent Tn-glycopeptide antigens on a cancer cell makes tumor escape due to antigen loss unlikely.

Topics: Adoptive Transfer; Animals; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Cell Line; Glycosylation; Humans; Membrane Glycoproteins; Mice; Neoplasms; Receptors, Chimeric Antigen

An impedimetric biosensor was developed for the selective detection of the cancer-associated T antigen, using the lectin from Arachis hypogaea (peanut agglutinin, PNA) as the recognition element. The increase in the biosensor's impedance after sample incubation was indicative of lectin recognition and complex formation between PNA and glycoproteins containing T antigen. When using asialofetuin as model glycoprotein, a minimum amount of 100 ng of glycoprotein could be detected, generating an increase in impedance of 7.2%. Albumin did not cause interference in the detection of T-carrying glycoproteins up to a concentration of 0.01 mg ml

Topics: Antigens, Tumor-Associated, Carbohydrate; Antigens, Viral, Tumor; Arachis; Biosensing Techniques; Equipment Design; Humans; Neoplasms; Peanut Agglutinin; Plant Lectins; Vicia

In the last two decades, the paramount importance of Tumor Associated Carbohydrate Antigens (TACAs) as targets for anticancer vaccine development has been firmly assessed. The Tn antigen is an ideal target for immunotherapy, in that it is masked on normal cells, but exposed on cancer cells. However, it is difficult to elicit an effective and long-lasting response against Tn antigen and other TACAs. Here we report on the Tn antigen analogues developed to boost the latent Tn immune response. Hopefully, the results reported herein will be of help for the rational design of effective TACA-based immunostimulants.

Topics: Adjuvants, Immunologic; Animals; Antigens, Tumor-Associated, Carbohydrate; Biomimetic Materials; Cancer Vaccines; Galactosides; Humans; Mice; Mice, Inbred BALB C; Neoplasms; Peptides, Cyclic; RAW 264.7 Cells

Utilizing natural antibodies to augment vaccine immunogenicity is a promising approach toward cancer immunotherapy. Anti-rhamnose (anti-Rha) antibodies are some of the most common natural anti-carbohydrate antibodies present in human serum. Therefore, rhamnose can be utilized as a targeting moiety for a rhamnose-containing vaccine to prepare an effective vaccine formulation. It was shown previously that anti-Rha antibody generated in mice binds effectively with Rha-conjugated vaccine and is picked up by antigen presenting cells (APCs) through stimulatory Fc receptors. This leads to the effective uptake and processing of antigen and eventually presentation by major histocompatibility complex (MHC) molecules. In this article, we show that natural human anti-Rha antibodies can also be used in a similar mechanism and immunogenicity can be enhanced by targeting Rha-conjugated antigens. In doing so, we have purified human anti-Rha antibodies from human serum using a rhamnose affinity column. In vitro, human anti-Rha antibodies are shown to enhance the uptake of a model antigen, Rha-ovalbumin (Rha-Ova), by APCs. In vivo, they improved the priming of CD4+ T cells to Rha-Ova in comparison to non-anti-Rha human antibodies. Additionally, increased priming of both CD4+ and CD8+ T cells toward the cancer antigen MUC1-Tn was observed in mice that received human anti-Rha antibodies prior to vaccination with a rhamnose-modified MUC1-Tn cancer vaccine. The vaccine conjugate contained Pam

Topics: Animals; Antibodies; Antigen-Presenting Cells; Antigens, Tumor-Associated, Carbohydrate; Cancer Vaccines; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Female; Humans; Immunogenicity, Vaccine; Mice; Mice, Inbred C57BL; Mucin-1; Neoplasms; Ovalbumin; Rhamnose

Tumor-associated carbohydrate antigens (TACAs) are attractive targets for anticancer vaccine development. Due to the low immunogenicity of TACAs, a powerful carrier system is needed to boost immune responses. Virus-like particles (VLPs) are an exciting platform for delivering TACAs to the immune system. The high symmetry of VLPs enables the display of TACAs in an organized manner, which in turn can potently activate antibody secreting B cells, eliciting high titers of antiglycan IgG antibodies. In this chapter, the protocol for conjugating a prototypical TACA, the Tn antigen to a VLP, bacteriophage Qβ, is presented. On an average around 370 copies of Tn can be attached to each Qβ capsid. Immunization of mice with Qβ-Tn conjugate leads to over two orders of magnitude higher IgG antibodies compared to control mice receiving Qβ only without the Tn antigen. Antibodies induced by Qβ-Tn recognize Tn-expressing tumor cells strongly and protect mice from tumor-induced death. The techniques for evaluating antibody titers by enzyme-linked immunosorbent assay, antibody binding to tumor cells by flow cytometry, and the protection efficacy of the vaccine in a therapeutic model of tumor are discussed in this chapter.

Topics: Allolevivirus; Animals; Antigens, Tumor-Associated, Carbohydrate; B-Lymphocytes; Biomarkers, Tumor; Disease Models, Animal; Flow Cytometry; Humans; Immunization; Mice; Neoplasms; Vaccines, Conjugate; Vaccines, Virus-Like Particle

Changes in glycosylation of the cancer cell glycocalyx are a hallmark of metastasizing cancers and critically contribute to distant metastasis. In this chapter we concentrate on two lectins capable of specifically binding tumor-associated glycans in cryostat or formalin-fixed, paraffin-embedded tissue sections derived from primary clinical material, genetically engineered mouse models with endogenous cancer formation or xenograft mouse models. The snail lectin of Helix pomatia (HPA) binds N-acetylgalactosamine (GalNAc) that is expressed among others as Tn antigen (O-linked GalNAc) in primary tumors and metastases in several human adenocarcinomas. Another lectin, Phaseolus vulgaris leucoagglutinin (PHA-L) binds to complex β1-6 branched N-linked oligosaccharides associated with increased metastatic potential in breast, colon, and prostate cancer. Using these two lectins both O- and N-linked alterations in the glycocalyx of cancer cells can be monitored. As they are commercially available in a biotinylated or fluorescence-labeled form they can be readily used in cancer metastasis studies.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Biomarkers; Cell Line, Tumor; Disease Models, Animal; Glycosylation; Heterografts; Histocytochemistry; Humans; Lectins; Mice; Microscopy; Neoplasm Metastasis; Neoplasms; Protein Binding

Legume lectins are the most thoroughly studied group of lectins and have been widely linked to many pathological processes. Their use as immunohistochemistry markers for cell profiling and cancer diagnosis have made these molecules important tools for immunological studies and have stimulated the prospection and characterization of new lectins. The crystal structures of a recombinant seed lectin from Vatairea macrocarpa (rVML) and its complexes with GalNAcα1-O-Ser, GalNAc and α-lactose, have been determined at 1.90, 1.97, 2.70 and 1.83Å resolution, respectively. Small angle X-ray scattering and calorimetry assays have confirmed the same pH stable oligomerization pattern and binding profiles proposed for its wild-type counterpart. In silico analyzes have explored the potential of this recombinant lectin as new tool for cancer research through a comparative profile with other legume lectins widely used for cancer diagnosis and prognosis. The results suggest the recognition of specific epitopes exhibited on different cancer cells as a process that relies on the disposition of hydrophobic clusters and charged regions around the lectin carbohydrate-binding site, favouring the anchorage of different groups in the antigen boundaries, highlighting the different potential of each analyzed lectin. In conclusion, the experimental results and comparative analysis show that rVML is as a promising tool for cancer research, able to bind with high affinity specific tumor-associated antigens, highly stable and easily produced.

Topics: Acetylgalactosamine; Antigens, Tumor-Associated, Carbohydrate; Fabaceae; Lactose; Molecular Docking Simulation; Neoplasms; Plant Lectins; Protein Binding; Protein Conformation

For successful carbohydrate based anti-cancer vaccines, it is critical that B cells are activated to secret antibodies targeting the tumor associated carbohydrate antigens (TACAs). Despite the availability of many TACA based constructs, systematic understanding of the effects of structural features on anti-glycan antibody responses is lacking. In this study, a series of defined synthetic glyco-polymers bearing a representative TACA, i.e., the Thomsen-nouveau (Tn) antigen, have been prepared to probe the induction of early B cell activation and antibody production via a T cell independent mechanism. Valency and density of the antigen in the polymers turned out to be critical. An average of greater than 6 Tn per chain was needed to induce antibody production. Glycopolymers with 40 antigens per chain and backbone molecular weight of 450 kDa gave the strongest stimulation to B cells in vitro, which correlated well with its in vivo activity. Deviations from the desired valency and density led to decreased antibody production or even antigen specific B cell non-responsiveness. These findings provide important insights on how to modulate anti-TACA immune responses facilitating the development of TACA based anti-cancer vaccines using glycopolymers.

Topics: Animals; Antibody Formation; Antigens, Tumor-Associated, Carbohydrate; B-Lymphocytes; Cancer Vaccines; Carbohydrates; Cells, Cultured; Female; Mice, Inbred C57BL; Neoplasms

Herein, we report a new process that enables the gram-scale production of a fully synthetic anti-cancer vaccine for human use. This therapeutic vaccine candidate, named MAG-Tn3, is a high-molecular-weight tetrameric glycopeptide encompassing carbohydrate tumor-associated Tn antigen clusters and peptidic CD4

Topics: Amino Acid Sequence; Animals; Antigens, Tumor-Associated, Carbohydrate; Cancer Vaccines; CD4-Positive T-Lymphocytes; Dendrimers; Glycopeptides; Humans; Mice; Mice, Transgenic; Neoplasms; Vaccines, Synthetic

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Cell Line, Tumor; Female; Humans; Immunization; Immunoglobulin G; Immunoglobulin M; Mice, Inbred C57BL; Mice, Transgenic; Mucins; Neoplasms; Programmed Cell Death 1 Receptor

Tumor-associated carbohydrate antigens (TACAs) are key components of cancer vaccines. A variety of vaccines based on native TACAs such as α-Tn have shown immunogenicity and protection in preclinical animal studies, however, their weak immunogenicity, low in vivo instability, and poor bioavailability, have discouraged their further evaluations in clinical studies. A new improved vaccine prototype is reported. It is composed of four clustered Tn-antigen mimetics and a immunogenic peptide epitope that are conjugated to a cyclopeptide carrier. The immunization of mice with this vaccine 1) was safe, 2) induced a strong and long-lasting Tn-specific response with IgM/IgG antibodies able to recognize native carbohydrate antigens; 3) produced high titers of IgG1, IgG2a, and IgG3 antibodies; and 4) produced a significant antibody-dependent regression of tumors and conferred protection. Altogether, these findings pave the way for the clinical development of safe and effective therapeutic vaccines against Tn-expressing cancers.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Cancer Vaccines; Humans; Mice; Neoplasms

The development of an effective immunotherapy is an attractive strategy toward cancer treatment. Tumor associated carbohydrate antigens (TACAs) are overexpressed on a variety of cancer cell surfaces, which present tempting targets for anticancer vaccine development. However, such carbohydrates are often poorly immunogenic. To overcome this challenge, we show here that the display of a very weak TACA, the monomeric Tn antigen, on bacteriophage Qβ virus-like particles elicits powerful humoral responses to the carbohydrate. The effects of adjuvants, antigen display pattern, and vaccine dose on the strength and subclasses of antibody responses were established. The local density of antigen rather than the total amount of antigen administered was found to be crucial for induction of high Tn-specific IgG titers. The ability to display antigens in an organized and high density manner is a key advantage of virus-like particles such as Qβ as vaccine carriers. Glycan microarray analysis showed that the antibodies generated were highly selective toward Tn antigens. Furthermore, Qβ elicited much higher levels of IgG antibodies than other types of virus-like particles, and the IgG antibodies produced reacted strongly with the native Tn antigens on human leukemia cells. Thus, Qβ presents a highly attractive platform for the development of carbohydrate-based anticancer vaccines.

Topics: Adjuvants, Immunologic; Animals; Antigens, Tumor-Associated, Carbohydrate; Bacteriophages; Cancer Vaccines; Capsid; Female; Humans; Immunity; Immunoglobulin G; Mice; Mice, Inbred C57BL; Models, Molecular; Neoplasms

The Tn antigen (GalNAcα-O-Ser/Thr) is a well-established tumor-associated marker which represents a good target for the design of anti-tumor vaccines. Several studies have established that the binding of some anti-Tn antibodies could be affected by the density of Tn determinant or/and by the amino acid residues neighboring O-glycosylation sites. In the present study, using synthetic Tn-based vaccines, we have generated a panel of anti-Tn monoclonal antibodies. Analysis of their binding to various synthetic glycopeptides, modifying the amino acid carrier of the GalNAc(*) (Ser* vs Thr*), showed subtle differences in their fine specificities. We found that the recognition of these glycopeptides by some of these MAbs was strongly affected by the Tn backbone, such as a S*S*S* specific MAb (15G9) which failed to recognize a S*T*T* or a T*T*T* structure. Different binding patterns of these antibodies were also observed in FACS and Western blot analysis using three human cancer cell lines (MCF-7, LS174T and Jurkat). Importantly, an immunohistochemical analysis of human tumors (72 breast cancer and 44 colon cancer) showed the existence of different recognition profiles among the five antibodies evaluated, demonstrating that the aglyconic part of the Tn structure (Ser vs Thr) plays a key role in the anti-Tn specificity for breast and colon cancer detection. This new structural feature of the Tn antigen could be of important clinical value, notably due to the increasing interest of this antigen in anticancer vaccine design as well as for the development of anti-Tn antibodies for in vivo diagnostic and therapeutic strategies.

Topics: Adult; Aged; Aged, 80 and over; Amino Acid Sequence; Animals; Antibodies, Monoclonal; Antibody Specificity; Antigens, Tumor-Associated, Carbohydrate; Biomarkers, Tumor; Breast Neoplasms; Cell Line, Tumor; Colonic Neoplasms; Female; Glycopeptides; Humans; Male; Mice; Middle Aged; Neoplasm Staging; Neoplasms; Protein Binding

Invasiveness underlies cancer aggressiveness and is a hallmark of malignancy. Most malignant tumors have elevated levels of Tn, an O-GalNAc glycan. Mechanisms underlying Tn up-regulation and its effects remain unclear. Here we show that Golgi-to-endoplasmic reticulum relocation of polypeptide N-acetylgalactosamine-transferases (GalNAc-Ts) drives high Tn levels in cancer cell lines and in 70% of malignant breast tumors. This process stimulates cell adhesion to the extracellular matrix, as well as migration and invasiveness. The GalNAc-Ts lectin domain, mediating high-density glycosylation, is critical for these effects. Interfering with the lectin domain function inhibited carcinoma cell migration in vitro and metastatic potential in mice. We also show that stimulation of cell migration is dependent on Tn-bearing proteins present in lamellipodia of migrating cells. Our findings suggest that relocation of GalNAc-Ts to the endoplasmic reticulum frequently occurs upon cancerous transformation to enhance tumor cell migration and invasiveness through modification of cell surface proteins.

Topics: Acetylgalactosamine; Animals; Antigens, Tumor-Associated, Carbohydrate; Blotting, Western; Cell Line; Cell Movement; Cloning, Molecular; Endoplasmic Reticulum; Fluorescent Antibody Technique; Gene Expression Regulation, Neoplastic; Glycosylation; Glycosyltransferases; Golgi Apparatus; Humans; Kaplan-Meier Estimate; Mice; Mice, Inbred BALB C; Neoplasm Invasiveness; Neoplasms

Mucin glycoproteins are major secreted or membrane-bound molecules that, in cancer, show modifications in both the mucin proteins expression and in the O-glycosylation profile, generating some of the most relevant tumour markers in clinical use for decades. Thus far, the identification of these biomarkers has been based on the detection of either the protein or the O-glycan modifications. We therefore aimed to identify the combined mucin and O-glycan features, that is, specific glycoforms, in an attempt to increase specificity of these cancer biomarkers. Using in situ proximity ligation assays (PLA) based on existing monoclonal antibodies directed to MUC1, MUC2, MUC5AC and MUC6 mucins and to cancer-associated carbohydrate antigens Tn, Sialyl-Tn (STn), T, Sialyl-Le(a) (SLe(a)) and Sialyl-Le(x) (SLe(x)) we screened a series of 28 mucinous adenocarcinomas from different locations (stomach, ampulla of Vater, colon, lung, breast and ovary) to detect specific mucin glycoforms. We detected Tn/STn/SLe(a)/SLe(x)-MUC1 and STn/SLe(a)/SLe(x)-MUC2 glycoforms in ≥50% of the cases, with a variable distribution among organs. Some new glycoforms-T/SLe(a)-MUC2, STn/T/SLe(a) SLe(x)-MUC5AC and STn/T/SLe(a)/SLe(x)-MUC6-were identified for the first time in the present study in a variable percentage of cases from different organs. In conclusion, application of the PLA technique allowed sensitive detection of specific aberrant mucin glycoforms in cancer, increasing specificity to the use of antibodies either to the mucin protein backbone or to the O-glycan haptens alone.

Topics: Adenocarcinoma, Mucinous; Antigens, Tumor-Associated, Carbohydrate; Biomarkers, Tumor; Breast; CA-19-9 Antigen; Colon; Fluorescent Antibody Technique; Gangliosides; Glycosylation; Humans; Immunohistochemistry; Lung; Mucin 5AC; Mucin-1; Mucin-2; Mucin-6; Neoplasms; Oligosaccharides; Sialyl Lewis X Antigen

Topics: Antigens, Tumor-Associated, Carbohydrate; Antigens, Viral, Tumor; Binding Sites; Carbohydrate Conformation; Carbohydrate Sequence; Crystallography, X-Ray; Epitopes; Humans; Models, Molecular; Molecular Sequence Data; Neoplasms; Oligosaccharides; Plant Lectins; Plants; Protein Binding

Oncogenic antigens such as Tn-antigen (GalNAcα-Ser/Thr) are involved in metastatic processes and are associated with a poor prognosis, thus representing excellent targets for cancer intervention. Available anti-Tn antibodies which can be applied for therapeutics or diagnostics are severely limited mostly because the Tn-antigen epitope by itself is too small to be antigenic in addition to the fact that many carbohydrates are self-antigens. To characterize anti-Tn monoclonal antibodies as well as to perform panning and screening for isolation of anti-Tn single chain variable fragments from phage-display libraries, a large quantity of inexpensive Tn-antigens are needed. In this study, thus, glycophorin A which is a highly glycosylated sialoglycoprotein with approximately 12 O-glycans was sequentially treated with sialidase and β-galactosidase to remove sialic acid and galactose residues. The resulted product was shown to be an asialo-agalacto-glycophorin A which is reactive to an anti-Tn-antigen antibody. The simple preparation procedures described here would greatly help production and characterization of potentially valuable anti-Tn-antigen antibodies, which can be readily developed for cancer therapeutics and diagnostics.

Topics: Antibodies; Antibodies, Monoclonal; Antigens, Tumor-Associated, Carbohydrate; beta-Galactosidase; Enzyme-Linked Immunosorbent Assay; Epitopes; Glycophorins; Humans; N-Acetylneuraminic Acid; Neoplasm Metastasis; Neoplasms; Neuraminidase; Oligosaccharides; Peptide Library; Polysaccharides

To evaluate the differences in glycoforms of ascitic fluids which can be considered as useful in discrimination between benign and malignant patients.. The investigations were carried out on 17 benign and 13 malignant patients. To obtain high molecular weight material, gel exclusion chromatography was applied. To evaluate the relative amounts of different glycoforms, ELISA tests with biotinylated lectins were used.. The fucosylation pattern was found to be similar in malignant and benign group. Fucose linked by alpha 1-6 bond was the most abundant structure. Sialylation was found to be more typical for malignant patients. Alpha 2-6 bond was observed on higher level than alpha 2-3 linkage. T and Tn antigens were present on rather low level with a slight prevalence of T antigen in malignant group. The glycoforms recognized by DSA lectin were more numerous in benign patients.. The evaluation of the level of some ascitic fluids glycoforms can be useful in differentiation between malignant and benign diseases.

Topics: Antigens, Tumor-Associated, Carbohydrate; Antigens, Viral, Tumor; Ascitic Fluid; Enzyme-Linked Immunosorbent Assay; Female; Fucose; Glycosylation; Humans; Lectins; Male; Mucin-1; Neoplasms

Malignant transformation of epithelial cells is frequently associated with the alteration of glycosylation pathways. Tn is a common tumor-associated carbohydrate antigen present in 90% of human carcinomas and its expression correlates with metastatic potential and poor prognosis. Despite its relevance, the functional role of Tn in tumor biology has not been firmly established probably for the lack of appropriate experimental tools. Our aims were to produce highly reactive monoclonal antibodies against Tn making use of synthetically produced Tn and to test their usefulness for in vivo imaging as well as to define their potential functional activity in tumor cell spread. We immunized mice with Tn clustered on cationized BSA and screened the positive hybridomas with Tn-biotinylated alginate. Enzyme-linked immuno sorbent assay and immunofluorescence assays revealed that the most reactive anti-Tn IgM mAb (2154F12A4) selectively recognized Tn on the MCF7 breast cancer cell line since its binding to the cell membrane was completely abolished by preincubation with purified Tn. Importantly, QDot 800-conjugated mAb injected in MCF7-tumor bearing mice specifically bound to primary tumor lesions as well as to metastases in lymph nodes. In addition, this mAb was able to inhibit cancer cell adhesion to lymphatic endothelium suggesting a novel involvement of Tn in the lymphatic dissemination of cancer cells and hypothesizing future applications in inhibiting lymphatic metastases.

Topics: Animals; Antibodies, Monoclonal; Antibodies, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Cell Adhesion; Cell Line, Tumor; Endothelium; Female; Humans; Immunohistochemistry; Lymph Nodes; Lymphatic Metastasis; Mice; Mice, Inbred BALB C; Mice, Nude; Molecular Structure; Neoplasms; Xenograft Model Antitumor Assays

An entirely carbohydrate-based immunogen consisting of a zwitterionic polysaccharide (ZPS) PS A1 and the well-known tumor antigen Tn has been designed, synthesized, and studied for immunological effects. The PS A1 motif was included to act as an MHCII elicitor for a T-cell-dependent immune response with increased immunogenicity against tumor-associated carbohydrate antigens, providing an alternative to carrier proteins. Through the use of C57BL/6 mice, it has been shown that chemical modification of PS A1 does not alter the recognition sequence responsible for an MHCII-mediated, T-cell-dependent immune response. The Tn-PS A1 conjugate construct confers specificity toward the Tn antigen alone, and specific carbohydrate immunoglobulins, namely, IgG3, are generated from intraperitoneal immunizations with or without adjuvant. The properties of the vaccine candidate are attributed to a site-specific linking strategy that incurs significant incorporation of Tn antigen.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Cattle; Haptens; Histocompatibility Antigens; Immunoconjugates; Immunotherapy; Mice; Neoplasms; Oligosaccharides; Vaccines, Synthetic

Tn-antigens are generally masked by covalently linked carbohydrates but are exposed in most primary and metastatic epithelial malignant tumors, providing sensitive markers for detection of carcinoma. Here, therapeutic potentials of tumor-associated carbohydrate antigen-specific antibodies were investigated. MLS128, an anti-Tn monoclonal antibody, binds to a carbohydrate epitope consisting of three consecutive Tn-antigens (GalNAcalpha-Ser/Thr). MLS128 treatment significantly inhibited colon and breast cancer cell growth. MLS128 bound to 110-210 kDa glycoproteins on the cell surface. MLS128 treatment caused down-regulation of insulin-like growth factor-I receptor and epidermal growth factor receptor in LS180 colon cancer cells, suggesting that MLS128-inhibited cancer cell growth is in part mediated by down-regulation of growth factor receptors. This study provides the first insights into the potential use of this particular type of anti-Tn antigen antibodies as a cancer therapeutic.

Topics: Antibodies, Monoclonal; Antigens, Tumor-Associated, Carbohydrate; Blotting, Western; Cell Line, Tumor; Cell Proliferation; Humans; Neoplasms

A novel anticancer vaccine candidate built on a nonpeptidic scaffold has been synthesized. Four S-Tn tumor-associated glycomimetic antigens have been clustered onto a calix[4]arene scaffold bearing an immunoadjuvant moiety (P3CS). The immunogenicity of the synthetic construct has been investigated by immunization of mice in vivo. ELISA assay has evidenced that the tetravalent construct stimulates a higher production of anti-Tn antigen IgG antibodies when compared to an analogous monovalent compound. This result is ascribable to an antigen cluster effect and makes the reported vaccine candidate a good mimic of the natural motifs present on the mucine surface.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Calixarenes; Cancer Vaccines; Chromatography, Thin Layer; Dipeptides; Enzyme-Linked Immunosorbent Assay; Female; Immunoglobulin G; Immunotherapy; Indicators and Reagents; Lipoproteins; Magnetic Resonance Spectroscopy; Mice; Mice, Inbred BALB C; Molecular Mimicry; Neoplasms; Phenols; Spectrometry, Mass, Electrospray Ionization

Neoplastic lesions typically express specific carbohydrate antigens on glycolipids, mucins, and other glycoproteins. Such antigens are often under epigenetic control and are subject to reversion and loss upon therapeutic selective pressure. We report here that two of the most common tumor-associated carbohydrate antigens, Tn and sialyl Tn (STn), result from somatic mutations in the gene Cosmc that encodes a molecular chaperone required for formation of the active T-synthase. Diverse neoplastic lesions, including colon cancer and melanoma-derived cells lines, expressed both Tn and STn antigen due to loss-of-function mutations in Cosmc. In addition, two human cervical cancer specimens that showed expression of the Tn/STn antigens were also found to have mutations in Cosmc and loss of heterozygosity for the cross-linked Cosmc locus. This is the first example of somatic mutations in multiple types of cancers that cause global alterations in cell surface carbohydrate antigen expression.

Topics: Antigens, Tumor-Associated, Carbohydrate; Cell Line, Tumor; Colorectal Neoplasms; Female; Galactosyltransferases; Humans; Jurkat Cells; Melanoma; Molecular Chaperones; Neoplasms; Transfection; Uterine Cervical Neoplasms

Vicia villosa B(4) (VVL-B(4)) is an important lectin for detecting exposed Tn (GalNAcalpha1-Ser/Thr) determinants on cancer cells. In order to elucidate the binding factors involved in VVL-B(4) and glycotope interaction, the binding properties of this lectin were analyzed by enzyme-linked lectinosorbent and inhibition assays. From the results, it is concluded that the most critical factor affecting VVL-B(4) binding is polyvalency at the alpha anomer of Gal with -NH CH(3)CO at carbon-2 (Tn epitope), which enhances the reactivity by 3.3x10(5) times over monovalent Gal. The reactivities of glycotopes can be ranked as follows: high density Tn cluster >>Tn glycopeptides (MW<3.0x10(3) >> monomeric Tn to tri- Tn glycopeptides >>> other GalNAcalpha/beta-related structural units>Gal and Galalpha- or beta-linked ligands, demonstrating the essential role of the polyvalency of Tn glycotopes in the enhancement of the binding.

Topics: Animals; Antigens, Tumor-Associated, Carbohydrate; Binding Sites; Carbohydrate Sequence; Glycoproteins; Humans; N-Acetylneuraminic Acid; Neoplasms; Oligosaccharides; Plant Lectins; Vicia

Blood group-related antigens have been attractive targets for immunotherapy of cancer since their initial identification as cancer-related antigens. However, available information on the relative expression of most of these antigens on human malignant and normal tissues has been insufficient for selecting optimal antigens and tumors for immune attack. In this study, the distribution of the blood group-related antigens TF, Tn, sTn, Le(a), sialyl Le(a), Le(b), Le(x), sialyl Le(x), polyfucosyl Le(x) and Le(y) on 13 types of cancer and 16 normal tissues was compared. Our results show that sTn is strongly expressed on cancers of breast, colon, stomach, ovary, prostate and uterus; Tn on prostate cancer; TF on cancers of breast, colon, ovary, prostate and uterus; Le(y) on the cancers of colon, lung, pancreas and ovary; Le(a) and Le(x) on gastric cancer; and sialyl Le(a) and sialyl Le(x) on colon cancer. The complete absence of these antigens on cancers of neuroectodermal or mesodermal origin including melanoma, sarcoma, neuroblastoma and B cell lymphoma is as striking as their widespread presence on tumors of epithelial origin. Normal tissues were also tested. Tn and Le(b) were only detected on gastric and ovarian epithelia; sTn on Leydig cells of testis in addition to gastric and ovarian epithelia; Le(x) and sialyl Le(x) on polymorphonuclear leukocytes; and TF, Le(a), sialyl Le(a), Le(x), sialyl Le(x), polyfucosyl Le(x) and Le(y) on epithelia from a variety of tissues.

Topics: Animals; Antibodies, Monoclonal; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Humans; Immunohistochemistry; Lewis Blood Group Antigens; Mice; Neoplasms

In the treatment of infections arisen in neoplastic patients without neutropenia, 2 antibiotic combinations (aztreonam + oxacillin vs tobramycin + cefoxitin), have been compared with regard to therapeutic effectiveness and tolerability. Twenty patients (age: 30-75) have been studied. Tolerability of both combinations was excellent. Results of this study showed a lower percentage of superinfections and a higher percentage of cure in patients treated with the combination aztreonam + oxacillin, even if the data were not statistically significant.

Topics: Adult; Aged; Aztreonam; Bacterial Infections; Cefoxitin; Drug Evaluation; Drug Therapy, Combination; Female; Humans; Male; Middle Aged; Neoplasms; Oxacillin; Tobramycin

Topics: Antibodies, Monoclonal; Antibodies, Neoplasm; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Bacteria; Cell Membrane; Enzyme-Linked Immunosorbent Assay; Epitopes; Erythrocyte Membrane; Hemagglutination Inhibition Tests; Humans; Intestines; Neoplasms

A monoclonal antibody, FBT3, was raised against Tn positive erythrocytes and, using immunohistochemistry, fresh and fixed tissues from patients with cancer were studied to detect any expression of a Tn-like epitope. Expression was found in neoplastic cells, usually both in cytoplasm and on cell membranes, from 104 of 147 cases of carcinoma and 1 of 13 cases of lymphoma, but rarely in adjacent, morphologically normal cells. Tn expression was seen in some normal glandular cells but, unlike cancer cells, it was distributed as fine granules in a supranuclear position. Detection of a Tn-like epitope is of theoretical interest and may be of direct diagnostic value.

Topics: Animals; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Biomarkers, Tumor; Cell Membrane; Cytoplasm; Epitopes; Erythrocytes; Humans; Male; Mice; Mice, Inbred BALB C; Neoplasms

Synthetic carbohydrate haptens, which are conjugated to carrier human serum albumin molecules [synthetic tumor-associated glycoconjugates (S-TAGs)], were used to immunize mice for monoclonal antibody (MoAb) production. Two of the S-TAGs were composed of haptens related to the Thomsen-Friedenreich (TF) antigen, and their structures are beta Gal(1----3)-beta GalNAc (TF-beta) and beta Gal(1----3) alpha GalNAc (TF-alpha) (Gal = galactose; GaNAc = N-acetylgalactosamine). The third S-TAG was made up of Tn hapten groups of the structure alpha GalNAc-O-serine. MoAbs specific for TF-alpha and Tn were able to be generated. All MoAbs generated against TF-beta cross-reacted with TF-alpha but not with Tn. None of the TF-alpha-specific MoAbs reacted with human carcinomas, whereas several TF-beta and Tn MoAbs were found to react with most human lung, colon, and breast carcinomas. It is believed that this is the first report of the use of synthetic carbohydrate cancer antigens for the production of anticancer MoAbs.

Topics: Antibodies, Monoclonal; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Disaccharides; Haptens; Humans; Neoplasms; Serum Albumin

Primary and metastatic carcinomas are epithelial in origin and comprise by far the largest group of malignant tumors in humans. In most of these tumors, T and Tn antigens, whose epitopes have been synthesized, are uncovered and immunoreactive. In all other tissues T and Tn antigens are masked and not accessible to the immune system; they are generally precursors in normal complex carbohydrate chains. Thus, carcinomas have antigens recognized as foreign by the patients' immune system. The expression of T and Tn antigens has pathogenic and clinical consequences, and the antigens themselves are powerful histological markers in carcinoma diagnosis and frequently in prognosis. Most patients distinguish their carcinoma from all other cells, as shown by strong autoimmune responses to T antigen. These responses are readily measured by assays, and they allow detection of carcinomas with greater sensitivity and specificity frequently earlier than previously possible. Moreover, the extent of T and Tn expression often correlates with carcinoma differentiation; on a molecular level, clustered T- and Tn-active structures on carcinoma cell surfaces may be involved in invasion.

Topics: Antibody Formation; Antigens; Antigens, Neoplasm; Antigens, Tumor-Associated, Carbohydrate; Antigens, Viral, Tumor; Autoantigens; Breast Neoplasms; Cell Adhesion; Humans; Immunity, Cellular; Lung Neoplasms; Neoplasm Metastasis; Neoplasms