A calicheamicin conjugate with a fully humanized anti-MUC1 antibody shows potent antitumor effects in breast and ovarian tumor xenografts.

Murine CTM01 is an internalizing murine IgG(1) monoclonal antibody that recognizes the MUC1 antigen expressed on many solid tumors of epithelial origin. Calicheamicin conjugates of this antibody have previously been shown to be potent, selective antitumor agents in preclinical models. A conjugate has now been made with a genetically engineered human version of this antibody, hCTM01.

CPI-0004Na, a new extracellularly tumor-activated prodrug of doxorubicin: in vivo toxicity, activity, and tissue distribution confirm tumor cell selectivity.

The search for cancer therapies that are more selective for tumor cells and spare normal sensitive cells has been very active for at least 20 years. The extracellularly tumor-activated peptidic prodrug of doxorubicin (Dox) CPI-0004Na (N-succinyl-beta-alanyl-L-leucyl-L-alanyl-L-leucyl-Dox) is potentially such a treatment. Here, we report the results of lethality studies performed with this compound in the mouse, showing that it is up to 4.

Gemtuzumab ozogamicin, a potent and selective anti-CD33 antibody-calicheamicin conjugate for treatment of acute myeloid leukemia.

CD33 is expressed by acute myeloid leukemia (AML) cells in >80% of patients but not by normal hematopoietic stem cells, suggesting that elimination of CD33(+) cells may be therapeutically beneficial. A conjugate of a calicheamicin hydrazide derivative attached via hydrazone formation to the oxidized carbohydrates of the anti-CD33 murine antibody P67.6 had been chosen for use in AML prior to humanization of this antibody.

N-Succinyl-(beta-alanyl-L-leucyl-L-alanyl-L-leucyl)doxorubicin: an extracellularly tumor-activated prodrug devoid of intravenous acute toxicity.

Intravenous administration of N-(beta-alanyl-L-leucyl-L-alanyl-L-leucyl)doxorubicin (4) induces an acute toxic reaction, killing animals in a few minutes. This results from its positive charge at physiological pH combined with its propensity to form large aggregates in aqueous solutions. Negatively charged N-capped versions of 4 such as the succinyl derivative 5 can be administered by the iv route at more than 10 times the LD(50) of doxorubicin without inducing the acute toxic reaction, and they are active in vivo.

Extracellularly tumor-activated prodrugs for the selective chemotherapy of cancer: application to doxorubicin and preliminary in vitro and in vivo studies.

Oligopeptidic derivatives of anthracyclines unable to penetrate cells were prepared and screened for their stability in human blood and their reactivation by peptidases secreted by cancer cells. N-beta-alanyl-L-leucyl-L-alanyl-L-leucyl-doxorubicin was selected as a new candidate prodrug. The NH2-terminal beta-alanine allows a very good blood stability.

Radioimmunotherapy of colorectal carcinoma xenografts in nude mice with yttrium-90 A33 IgG and Tri-Fab (TFM).

The monoclonal antibody A33 recognises a tumour-associated antigen on human colorectal carcinoma, and has undergone preliminary evaluation in the clinic where selective localisation to hepatic metastases has been demonstrated [Welt et al. (1994) J. Clin.

Comparative metabolism and retention of iodine-125, yttrium-90, and indium-111 radioimmunoconjugates by cancer cells.

Radiolabeled antibodies have produced encouraging remissions in patients with chemotherapy-resistant hematological malignancies; however, the selection of therapeutic radionuclides for clinical trials remains controversial. In this study, we compared the internalization, lysosomal targeting, metabolism, and cellular retention of radiolabeled murine and humanized monoclonal antibodies targeting the CD33 antigen (monoclonal antibodies mP67 and hP67, respectively) on myeloid leukemia cell lines (HEL and HL-60) and of anti-carcinoma antibodies (monoclonal antibodies hCTM01 and hA33) targeting breast cancer and colorectal carcinoma cell lines (MCF7 and Colo 205, respectively). Each antibody was labeled with 125I (by the IodoGen method) and with 111In and 90Y using macrocyclic chelation technology.

Pharmacokinetics and scintigraphy of indium-111-DTPA-MOC-31 in small-cell lung carcinoma.

Unlabelled: Radiolabeled MOC-31 retains its immunoreactivity and shows good in vivo immunolocalization to human SCLC xenografted in nude rats.

Methods: We evaluated the immunotargeting properties and safety of 111In-labeled monoclonal antibody (MAb) MOC-31 (125 MBq, 5 mg) in six patients with histologically proven small-cell lung cancer (SCLC). Scintigraphy and pharmacokinetics were performed up to 3 days after injection.

Potentiation of antiproliferative effects of monoclonal antibody Lym-1 and immunoconjugate Lym-1-gelonin on human Burkitt's lymphoma cells with gamma-interferon and tumor necrosis factor.

A type I ribosome inactivating protein, gelonin, was linked to Lym-1, a murine monoclonal antibody reactive with a polymorphic determinant of class II HLA-DR histocompatibility leukocyte antigen (HLA) on human lymphoma cells, via a disulfide linkage using the heterobifunctional cross-linking agent, N-succinimidyl-3-(2-pyridyldithio) propionate. This immunotoxin was purified from unreacted gelonin and unconjugated Lym-1 by fast protein liquid chromatography using sephacryl S-300 gel filtration and blue sepharose affinity gradient separation. Binding of Lym-1-gelonin immunoconjugate to human Raji Burkitt's lymphoma cells was demonstrated by indirect immunofluorescence using flow cytometry.

A phase I escalating-dose safety, dosimetry and efficacy study of radiolabeled monoclonal antibody LYM-1.

Thirteen patients with relapsed or refractory Non-Hodgkin's Lymphoma were treated with 131I-Lym-1 during the course of a dose escalation trial. Principal aims were to establish the maximum tolerated single dose (MTD), as well as to assess clinical and dosimetric effects of the MTD. Patients were eligible if > 25% of tumor cells bound Lym-1 on immunohistochemistry, stain intensity was +2/4 or greater and human anti-mouse antibody (HAMA) assay was negative.

Dose fractionation of radiolabeled antibodies in patients with metastatic colon cancer.

Twelve patients with metastatic colon cancer were treated with 131I-chimeric B72.3 (IgG-4) at total doses of 28 or 36 mCi/m2 in two or three weekly fractions. Bone marrow suppression was the only significant side effect.

Studies on the structure of the organ-specific determinant of human colonic mucin.

We had previously shown that the human colon produces at least two immunochemically distinct mucins, one neutral and the other a sialomucin [Gold et al. J. biol.

Comparison of tumor targeting of mouse monoclonal and goat polyclonal antibodies to carcinoembryonic antigen in the GW-39 human tumor-hamster host model.

We have evaluated 4 radioiodinated mouse monoclonal anticarcinoembryonic antigen antibodies (MAbs) by using the GW-39 human colorectal tumor xenograft transplanted i.m. in immunocompetent hamsters to determine whether there were any differences in their tumor localization properties.

Purification and characterization of carcinoembryonic antigen from GW-39, a xenografted human colonic tumor system.

Carcinoembryonic antigen (CEA) was purified from GW-39 human tumor xenografts in hamsters by immunoaffinity chromatography. Binding of the antigen to immobilized monoclonal antibody provided a high degree of purification of CEA in a single step. A recovery of 79% and a 750-fold purification were obtained.

Representation of epitopes on colon-specific antigen-p defined by monoclonal antibodies.

Colon-specific antigen-p (CSAp) is a large molecular-sized protein restricted to normal and neoplastic gastrointestinal tissues and to some mucinous ovarian tumors. Murine monoclonal antibodies (MAbs) were raised against CSAp that was affinity purified with goat polyclonal antibodies from GW-39 human colonic carcinoma xenografts or against the CSAp-producing colon cancer cell line SW-948. Two of the MAbs, designated Mu-2 and Mu-4, recognized a CSAp determinant containing sialic acid, and this epitope was also expressed on bovine submaxillary mucin (BSM).

Imaging of primary and metastatic liver cancer with 131I monoclonal and polyclonal antibodies against alphafetoprotein.

Thirteen patients with a history of confirmed liver carcinoma were given either I131 goat polyclonal or murine monoclonal antibodies against alpha-fetoprotein (AFP), and then scanned with a gamma camera. In order to reduce background, nontarget activity, especially in the liver, blood pool, and reticuloendothelial system, 99mTc imaging agents were used for tumor image enhancement by computer-assisted subtraction. A sensitivity of 91% for the primary site, 50% for the lungs (33% for the chest area), and 75% for the abdomen and pelvis was achieved, with a specificity of 100%, 94%, and 100% for these sites, respectively.

The identification of epitopic sites in human alpha 1-proteinase inhibitor.

Human alpha 1-proteinase inhibitor (alpha 1-PI) yielded nine fragments on cleavage with CNBr. The amino acid sequences of these fragments were determined. Three of these CNBr-cleavage fragments, namely fragment I (residues 64-220), fragment II (residues 243-351) and fragment III (residues 1-63), were found to bind rabbit polyclonal antibodies against chemically oxidized alpha 1-PI and mouse polyclonal antibodies against native alpha 1-PI by the Bio-Dot method (enzyme-linked immunosorbent assay on nitrocellulose).

Organ specific antigens of the human gastrointestinal tract.

We previously reported the characterization of a normal adult colonic mucin antigen which contained an organ specific immunodeterminant [Tissue Antigen 11, 362 (1978)]. In the present study we have investigated mucins produced at other levels of the gastrointestinal tract in order to determine if regional specificities exist. Mucins were isolated from normal adult stomach, jejunum, ileum and colon and used to prepare antisera in rabbits.

Characterization of two metastatic subpopulations originating from a single human colon carcinoma.

Two separate cell lines originating from distinct metastatic deposits in a patient with a primary colonic carcinoma have been established both in vivo and in vitro. One metastasis, OM-1, was found in the omentum, and the other, HOT-3, was located on the ovary. These two metastases differ in several significant characteristics, including growth kinetics and the production of carcinoembryonic antigen by the cultured cells.

Differential expression of tumor-associated antigens in human colon carcinomas xenografted into nude mice.

The tumor-associated antigen profile of a number of different human colon carcinomas xenografted into inbred Swiss nude mice was examined to determine whether the tumors could serve as useful models for antigen purification, radioimmunodetection, and immunotherapy. Extreme heterogeneity was observed both by radioimmunoassay and immunohistochemical procedures for the expression of carcinoembryonic antigen (CEA), colon-specific antigens (CSAp), and colonic mucin antigen (CMA) within the tumors. Four of 10 tumors (DLD-2, DLD-3, DLD-5, and HCT-10) were high producers of CEA (greater than 75 micrograms/g wet tissue wt).

Colon-specific antigen-p (CSAp). II. Further characterization in colorectal and pancreatic cancer.

The physicochemical and immunological characteristics of colon-specific antigen-p (CSAp) in plasma and in colorectal and pancreatic tumors were investigated. CSAp in the plasma of a rectal cancer patient and in a colonic carcinoma xenografted in hamsters (GW-39 tumor) appeared to have similar chromatographic properties, being of a molecular size of 4 million or more. The activities of CSAp in both plasma and tumor were similarly destroyed by treatment with a thiol reagent.

Colon-specific antigen-p (CSAp). I. Initial clinical evaluation as a marker for colorectal cancer.

A radiometric immunoassay for detecting colon-specific antigen-p (CSAp) in the blood of patients suspected of having colorectal cancer has been developed and evaluated in 272 subjects of various disease entities. Using 10 units/ml as the cutoff value for normalcy, the results indicate that the highest number of elevated CSAp titers occurred in patients with advanced colorectal cancer (61%). Only one of 12 colonic adenoma patients had an elevated CSAp titer, and this was slightly above the 10 units/ml cutoff.

alpha 1-Proteinase inhibitor production by human adenocarcinomas xenotransplanted into nude mice.

Human colon adenocarcinomas xenotransplanted into BALB/c nude mice were examined for the ectopic synthesis of alpha 1-proteinase inhibitor(s) (alpha 1Pi). Tumor extracts were prepared by homogenization followed by molecular sieve chromatography on Sepharose 4B. Fractions with immunoreactive alpha 1Pi were concentrated and adsorbed onto a concanavalin A-Sepharose 4B affinity column, then eluted, concentrated, and analyzed.

Protease digestion of colonic mucin. Evidence for the existence of two immunochemically distinct mucins.

Colonic mucin was prepared by phenol-water partition extraction of th colonic mucosa, followed by ethanol precipitation of the water-soluble material, molecular sieve chromatography, and reductive sodium dodecyl sulfate-disc gel electrophoresis in an agarose-polyacrylamide gradient gel. Although this material as observed to be relatively homogeneous by physical criteria, it was shown to contain at least two components by immunodiffusion. Molecular sieve, ion exchange, adsorption, lectin affinity, and electrophorectic techniques failed to separate the two components.

Characterization of colon-specific antigen-p and isolation of immunologically active tryptic peptides.

CSAp was originally detected with antisera to an unfractionated extract of GW-39 human colonic carcinoma xenografts, and was determined to be restricted to normal and neoplastic gastrointestinal tissues and certain ovarian tumors. Gel filtration chromatography of GW-39 extract on Sepharose 4B columns reveals that more than 90% of the CSAp is associated with the void volume fraction. The smaller m.