Therapeutic antibodies for prostate cancer.

Twenty five years ago, monoclonal antibodies were envisioned as magic bullets capable of targeting radioisotopes, toxins or cytotoxic drugs to the tumor site. It was soon realized that the potential of therapeutic antibodies far exceeded their use as carrier molecules and that native antibodies could also act as effector molecules capable of triggering a wide variety of antitumor responses. Today, we recognize that the utility and versatility of antibody-based products are unlimited; at the same time we have also learned that many obstacles need to be addressed to make antibody therapy an effective treatment modality.

Immune responses to murine monoclonal antibody-B43.13 correlate with prolonged survival of women with recurrent ovarian cancer.

Objective: We evaluated the therapeutic efficiency of the murine monoclonal antibody-B43.13 in the treatment of patients with recurrent ovarian cancer.

Study Design: This was a retrospective study of immune responses and survival in 44 patients who were treated with technetium 99m-labeled monoclonal antibody-B43.

Generation of CD4(+) and CD8(+) T lymphocyte responses by dendritic cells armed with PSA/anti-PSA (antigen/antibody) complexes.

Dendritic cells (DC) acquire antigens through a number of cell surface structures including receptors for the Fc portion of immunoglobulins and mannose. Little is known about the effects of antigen uptake via these receptors on antigen processing and presentation. We compared the capacity of DC to generate CD4(+) and CD8(+) T cell responses after exposure to prostate-specific antigen (PSA) alone, PSA targeted to the mannose receptor (mannosylated PSA (PSA-m)), or PSA targeted to Fc receptors by combining PSA with an anti-PSA antibody (AR47.

Induction of CA125-specific B and T cell responses in patients injected with MAb-B43.13--evidence for antibody-mediated antigen-processing and presentation of CA125 in vivo.

The murine monoclonal anti-CA125 antibody MAb-B43.13 has previously been administered as an immunoscintigraphic agent in order to monitor recurrence of ovarian cancer in patients, and a long-term follow-up demonstrated a survival benefit for these patients. The clinical benefit was initially attributed to the activation of the idiotypic network.

The effects of circulating antigen on the pharmacokinetics and radioimmunoscintigraphic properties of 99m Tc labelled monoclonal antibodies in cancer patients.

Unlabelled: PURPOSE. This article reports the pharmacokinetics, radiation dosimetry and radioimmunoscintigraphy (RIS) of two (99m)Tc-labelled monoclonal antibodies (MAb) used to detect cancer.

Methods: The effects of circulating antigen in female cancer patients are explored and their effects on the ability of these MAbs to effectively perform as RIS agents noted.

Immunotherapy of human ovarian carcinoma with OvaRex MAb-B43.13 in a human-PBL-SCID/BG mouse model.

The monoclonal antibody (MAb) B43.13, binding to the ovarian cancer-associated antigen CA125, has been injected into more than 200 patients with ovarian cancer to detect recurrence of the disease. The follow-up of the patients revealed surprisingly long survival spans for several patients despite high CA125 levels.

Expression of a fusion protein of scFv-biotin mimetic peptide for immunoassay.

We constructed two fusion proteins of scFv linked to biotin mimetic sequence (BMS) via different linkers, and expressed them in the Pichia pastoris expression/secretion system. We found that both bi-functional scFv proteins exhibited their intrinsic binding activities to antigen CA125 determined in competitive radioimmunoassay experiments, but the fusion protein with a spacer between the scFv and BMS (scFv-spacer-BMS) showed higher binding activity of streptavidin than the one with c-Myc peptide as a linker.

Use of encapsulated single chain antibodies for induction of anti-idiotypic humoral and cellular immune responses.

The use of biodegradable poly(lactic-co-glycolic acid) (PLGA) microspheres as a cancer vaccine delivery system for induction of anti-idiotypic responses has been investigated using a single chain antibody scFv-pDL10, which recognizes the human ovarian cancer antigen CA125. Immunization of mice with scFv-pDL10 encapsulated in PLGA microspheres resulted in enhanced humoral and cellular immune responses when compared to scFv-pDL10 alone. Induced anti-idiotypic antibodies (Ab2) which mimic the original antigen CA125 compete with CA125 for the epitope.

Induction of anti-idiotypic humoral and cellular immune responses by a murine monoclonal antibody recognizing the ovarian carcinoma antigen CA125 encapsulated in biodegradable microspheres.

The use of biodegradable poly(DL-lactic-co-glycolic acid) microspheres as a cancer vaccine delivery system for induction of anti-idiotypic responses was investigated using a murine monoclonal antibody B43.13 that recognizes the human ovarian cancer antigen CA125. Immunization of mice with mAb B43.

Summary report on the ISOBM TD-6 workshop: analysis of 20 monoclonal antibodies against Sialyl Lewisa and related antigens. Montreux, Switzerland, September 19-24, 1997.

The ISOBM TD-6 Workshop is the first international workshop on monoclonal antibodies against the Sialyl Lewisa (SLea) antigen. Eight research groups participated in a blind study to characterize the epitope binding, relative affinity and performance in immunoradiometric assays, of a panel of 20 monoclonal antibodies. The antibodies were tested against a diverse panel of neoglycoconjugates, purified antigens and human serum pools from gastrointestinal malignancies.

Anti-idiotype induction therapy: anti-CA125 antibodies (Ab3) mediated tumor killing in patients treated with Ovarex mAb B43.13 (Ab1).

Intravenous injection of the murine monoclonal anti-CA125 antibody B43.13 (Ovarex: Ab1) into ovarian cancer patients led to the induction of an idiotypic network. Of the 75 patients who received one to ten injections of a 2-mg dose of the antibody, 48 developed anti-(mAb B43.

Pharmacokinetics and radiation dosimetry of 99Tcm-labelled monoclonal antibody B43.13 in ovarian cancer patients.

OVAREX MAb B43.13 is a new radiopharmaceutical based on a monoclonal antibody (MAb-B43.13) known to recognize CA 125, a tumour antigen associated with epithelial ovarian cancer.

Radiolabeling of monoclonal antibody B43.13 with rhenium-188 for immunoradiotherapy.

In this study we report a novel method for direct radiolabeling of monoclonal antibody B43.13 (MAb-B43.13) with 188Re and have evaluated the product's radiochemical, biochemical, immunochemical and selected biological properties.

An engineered bivalent single-chain antibody fragment that increases antigen binding activity.

Bivalent single chain Fv (scFv) was constructed by fusing a polypeptide extension containing one or two cysteines to the COOH-terminus of an scFv antibody fragment. The scFv protein was expressed and secreted in a recombinant Pichia pastoris system as a dimer with a C-terminal disulfide bridge, as determined by Western blot analysis under non-reducing conditions. We found that the scFv construct with one cysteine in the C-extension (scFv-1Cys) exhibited a much higher dimer/monomer ratio than the two cysteine counterpart (scFv-2Cys).

Role of transchelation in the uptake of 99mTc-MAb in liver and kidney.

High radioactivity in liver and kidney after administration of 99mTc-labeled antibodies is a major detriment to the use of radiolabeled antibodies for diagnosis and therapy. In the present study, the uptake mechanism of radioactivity by liver and kidney involving 99mTc moiety was investigated. The data of in vitro and in vivo thiol transchelation studies, biodistribution alteration of 99mTc-MAb after specific modulation of endogenous thiol containing compounds, and the finding of 99mTc-labeled cysteine and GSH in bile, urine and kidney after administration of 99mTc-MAb demonstrated that transchelation by thiols (cysteine and GSH) played an important role in the localization of radiotracer from 99mTc MAb in normal tissues such as liver and kidney.

OVAREX MAb-B43.13:IFN-gamma could improve the ovarian tumor cell sensitivity to CA125-specific allogenic cytotoxic T cells.

Various immunological parameters were studied in 100 ovarian cancer patients injected with the OVAREX therapeutic vaccine (the functional component of which is anti-CA125 MAb-B43.13) to explain the serendipitous observation of prolonged survival after such treatment. In addition to CA125-specific humoral and cellular responses, interferon-gamma (IFN-gamma) was also found to be induced in those patients receiving the vaccine.

Preparation and protein conjugation of a divinyl sulphone derivatized bifunctional chelating agent.

A new bifunctional chelating agent with a novel linking arm, 2-[p-¿N-benzyl-N-(2-vinylsulfoethyl)¿- (aminobenzyl)¿-1,3-propane-diamine-N,N,N',N'-tetraacetic acid (VS-PDTA) was synthesized and was conjugated to protein for the purpose of attaching radiometals to monoclonal antibodies (MAbs). The effect of various parameters such as ligand concentration, protein concentration, pH, temperature and reaction period on the conjugation have been examined using chromatographic (SE and TLC) analysis after labeling with 111In. The parameters and chemical variables studied have significant effects on the efficiency and rate of protein conjugation.

Studies on metabolism of directly labeled 99mTc-antibody in mice.

The elucidation on the metabolic products of the 99mTc-antibody conjugates may provide insights and approaches that would reduce the undesirable deposition of radioactive species in normal tissues. In this investigation, the radiolabeled species in blood, urine, bile and extracts of liver and kidney obtained at different times after the injection of a model antibody, 99mTc-MAb170, into mice were analyzed with various chromatographic methods. Ninety-nine to 100% of the radioactivity in serum was associated with intact MAb170.

Epitope mapping of prostate-specific antigen with monoclonal antibodies.

Prostate-specific antigen (PSA) is a widely used marker for screening and monitoring prostate cancer. We identified and characterized the epitopes of two anti-PSA monoclonal antibodies (mAbs) designated B80 and B87. The epitopes were initially mapped as nonoverlapping by developing a sandwich immunoassay to measure PSA with the two anti-PSA mAbs.

Generation of human monoclonal anti-idiotypic antibodies with specificity to the murine monoclonal anti-CA 125 antibody B43.13.

Two human monoclonal antibodies, HID-7E7 and ROB-6F2, were produced by EBV transformation of peripheral blood lymphocytes (PBL). PBL were obtained from a patient with ovarian cancer who had been exposed several times to a Tc-99m labeled murine monoclonal anti-CA 125 antibody (B43.13, Biomira, Edmonton) for immunoscintigraphy.

An antibody-lectin sandwich assay for the determination of CA125 antigen in ovarian cancer patients.

A two-step forward sandwich assay was developed for the determination of the ovarian tumour associated glycoconjugate antigen CA125 with anti-CA125 Monoclonal antibody B27.1 on the solid phase and 125I-labelled wheat germ lectin as tracer in the solution phase. This Mab-lectin heterosandwich assay was optimized and the clinical utility was evaluated in sera from healthy volunteers and ovarian cancer patients.

Studies on the uptake mechanisms of 99mTc-labeled antibodies by liver tissue.

In this study, the roles of various protein alterations on the uptake of 99mTc labeled antibodies by isolated hepatocytes and by selected tissues in vivo were explored. In vitro binding studies of radiolabeled antibodies with hepatocytes demonstrated that the uptake of the radiolabel was a function of incubation duration and dose dependent. The uptake of 99mTc-antibodies could be inhibited by excess unlabeled F(ab')2 and Fc fragments as well as intact antibody.

Synthesis and evaluation of a new bifunctional chelating agent for the preparation of radioimmunoconjugates.

The conjugation of radiometals to monoclonal antibodies results in agents for radioimmunoimaging and other medical applications. Due to remarkable ability to form stable metal complexes with a great number of metal ions in different oxidation states, polyaminocarboxylate chelates are useful tools for this purpose. Bifunctional chelators that can hold radiometals with high stability under physiological conditions are essential to avoid radiation damage to non-target organs.

A rapid non-selective method to generate quadromas by microelectrofusion.

A simple non-selective methodology was developed and standardized to generate desired hybrid-hybridoma or quadroma secreting bifunctional antibodies. This novel protocol is based on microelectrofusion on a meander chamber using a few hundred cells of each of the two parental hybridomas with no laborious drug selection procedures. Seeding approximately 10 cells per well in a 96-well microtitre plate after fusion in 200 microliters standard medium containing 20% FBS and 10% Origen growth factor generated positive quadromas secreting bispecific antibodies with good stability after the second reclone.

Direct labeling of monoclonal antibodies with technetium-99m by photoactivation.

Unlabelled: Direct radiolabeling methods currently rely on the addition of exogenous chemical reagents to create the necessary binding sites for 99mTc binding to monoclonal antibodies (MAbs). This work describes the use of ultraviolet (UV) light to facilitate photoactivation of MAbs for 99mTc radiolabeling.

Methods: The parameters of exposure wavelength and solution composition were investigated to provide a basis for further development.