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Biomedical subjects

B Ballou

Publications and source records attributed to B Ballou.

18 recordsLinked to original sources
PubMed1992

Tissue localization of methotrexate-monoclonal-IgM immunoconjugates: anti-SSEA-1 and MOPC 104E in mouse teratocarcinomas and normal tissues.

Methotrexate (MTX) was coupled to the tumor-targeting monoclonal IgM, anti-SSEA-1 and the non-targeting myeloma IgM, MOPC 104E. At 24-h intervals following injection, drug deposition in MH-15 teratocarcinomas and in several normal tissues was followed by immunoperoxidase microscopy using the M16 monoclonal antibody to MTX. MTX-anti-SSEA-1 was deposited on the surface and in the interior of living tumor cells 24 h after injection; at 48 h and after, only low-level binding to necrotic tissue was found. There was no significant gradation in staining from the outside to the interior of the tumors. In tumors, the control MOPC 104E immunoconjugate was detectable only in necrotic tissue. Binding to SSEA-1-expressing normal tissues was undetectable, except for pericryptal fibroblasts in the small intestine. No significant pathology was found in normal tissues that are SSEA-1 positive. High levels of the immunoconjugate were detected in the liver, where MTX was found predominantly in Kupffer cells and possibly in hepatocytes; again, no significant morphological changes were associated with this retention. Thus tumor-associated antigens can be suitable targets for antibody-drug conjugates even when present in normal tissues and in large quantities, provided that the antigens in normal tissues are inaccessible. Moreover, deposition in viable tumor tissue can be assessed using monoclonal antibodies to methotrexate.

Animals↗
PubMed1989

In vivo antitumor effect of methotrexate conjugated to a monoclonal IgM antibody specific for stage-specific embryonic antigen-1, on MH-15 mouse teratocarcinoma.

Methotrexate (MTX) was coupled to an IgM monoclonal antibody specific for stage-specific embryonic antigen-1 (SSEA-1), and the resulting immunoconjugate (MTX-anti-SSEA-1) was used for in vivo drug targeting in mice bearing MH-15 teratocarcinoma. Immunoconjugates having an average of 65 mol MTX/mol antibody retained full antigen-binding capacity. Mice bearing well-established tumors (approx. 1 g) were treated i.v. using the immunoconjugate. MTX-anti-SSEA-1 at 15 mg/kg of drug had significant antitumor activity with no significant systemic toxicity. Neither an irrelevant isotype-matched conjugate, MTX-MOPC-104E, prepared from the MOPC 104E myeloma protein, nor free MTX injected alone or with either antibody had any significant antitumor effect. These results indicate that IgMs can be effective drug carriers for tumor targeting in spite of their high molecular mass, and that antigens that are selectively accessible in tumors, even though present in normal tissues, can be suitable targets for in vivo chemoimmunotherapy.

Antibodies, Monoclonal↗
PubMed1988

Regulation of glucocorticoid receptors and Na-K ATPase activity by hydrocortisone in proximal tubular epithelial cells.

The effect of hydrocortisone (HC) in modulating glucocorticoid receptors (GR) and sodium-potassium adenosine triphosphatase (Na-K ATPase) activity was studied in primary cultures of immunoisolated murine proximal tubular epithelial cells (PTEC). Utilizing monoclonal antibody against stage-specific embryonic antigen-1, a homogeneous population of PTEC was obtained in high yield. The cells were cultured to confluence and further treated for 48 h in serum-free growth medium containing no HC (control); 50 nM HC; or 50 nM HC plus 20 nM of the antiglucocorticoid, RU 38486. PTEC treated with 50 nM HC had 56% of GR binding and 160% Na-K ATPase activity as compared to controls (P less than 0.01). GR binding was abolished by incubation in RU 38486 whereas Na-K ATPase fell below control values (P less than 0.05). Brief incubations of HC-treated PTEC with 0.5 mM ouabain resulted in a fall in GR binding without a change in Na-K ATPase activity. These data indicate that in PTEC, HC regulates GR binding and they suggest that stimulation of Na-K ATPase activity is a direct biological response to this receptor-hormone interaction. Thus, primary cultures of immunoaffinity-isolated PTEC offer a good model system for investigating the molecular basis underlying the regulation of GR binding and postreceptor events influenced by glucocorticoids.

Animals↗
PubMed1988

Isolation and partial characterization of melanoma-associated antigens identified by autologous antibody.

The study of the autologous immune response to cancer avoids the difficulties encountered in the use of xenoantisera and may identify antigens of physiological relevance. However, the low titer and incidence of autologous antibody to melanoma have hampered further evaluation. By utilizing acid dissociation and ultrafiltration of serum, we have been able to augment the detectable autologous immune response to melanoma in the majority of patients studied. In autologous system Y-Mel 84:420, serum S150 demonstrated a rise in titer from 1:32 in native sera to 1:262,044 after dissociation. The antigen detected by S150 was found to be broadly represented on melanoma, glioma, renal cell carcinoma, neuroblastoma, and head and neck carcinoma cell lines. It did not react with bladder or colon carcinoma, fetal fibroblasts, pooled platelets, lymphocytes and red blood cells, or autologous cultured lymphocytes. Using polyacrylamide gel electrophoresis, S150 detects a 66,000-mol wt antigen in spent tissue culture media and serum ultrafiltrate. In cell lysate two bands between 20,000 and 30,000 mol wt are detected by S150. The 66,000-mol wt antigen is sensitive to trypsin digestion and but is resistant to pepsin and heat inactivation. Exposure of spent media to trypsin results in the development of a 24,000-mol wt band that appears to correspond to the antigen detected in the cell lysate. The difference between the antigens detected in the cell lysate as compared with spent media and serum ultrafiltrate may be due to degradation during cell lysis. We conclude that melanoma-associated antigens are present in the serum of patients with melanoma and are shed or secreted by their tumor cells.

Antibodies, Neoplasm↗
PubMed1987

Radioimmunolocation of a heterotransplanted human choriocarcinoma (BeWo) using monoclonal anti-SSEA-1: pharmacokinetics.

Stage-Specific Embryonic Antigen-1 (SSEA-1), originally discovered on mouse teratocarcinomas, has since been found on some human non-seminomatous germ-cell tumors and adenocarcinomas, as well as on some adult mouse and human tissues. A monoclonal antibody to this antigen (anti-SSEA-1; IgM, kappa) was used for radioimmunolocation. Nude mice bearing the human choriocarcinoma BeWo, which is SSEA-1 positive, were injected using a mixture of [131I]anti-SSEA-1 and [125I]MOPC 104E, an unselected myeloma protein of the same heavy-chain isotype. Animals were sacrificed at 24 hour intervals; the radioactive deposition due to both antibodies was determined for both tumors and normal organs. Accumulation of anti-SSEA-1 in the tumor was consistantly rapid and specific, while little accumulation of the unselected myeloma protein occurred. At five days after injection, an average of 3% of the initial dose of specific antibody was retained per gram of tumor; the tumor/blood ratio was 11, tumor/muscle was 80. Gamma-camera imaging allowed ready location of the tumors. Tumors could also be imaged using F(ab')2 antibody fragments.

Animals↗
PubMed1986

Tumor location and drug targeting using a monoclonal antibody (anti-SSEA-1) and antigen-binding fragments.

Both murine and heterotransplanted human nonseminomatous germ-cell tumors have been successfully located by external scintigraphy using radioiodinated anti-SSEA-1, a monoclonal IgM, and its pepsin-derived antigen-binding fragment, F(ab')2 mu. Antibody localization in the tumor is mainly due to antigenic specificity, rather than to nonspecific trapping, and depends strongly on the amount of time after injection. The antibody has been used for drug targeting in vitro and in vivo.

Animals↗
PubMed1986

Targeting, internalization, and cytotoxicity of methotrexate-monoclonal anti-stage-specific embryonic antigen-1 antibody conjugates in cultured F-9 teratocarcinoma cells.

Methotrexate (MTX) conjugates of a monoclonal antibody, anti-SSEA-1, containing an average of 45 mol MTX/mol of immunoglobulin M, were prepared by a carbodiimide coupling reaction. Binding experiments indicate that conjugation does not decrease the affinity of the antibody for its antigen. The conjugate strongly inhibits the growth of SSEA-1-bearing F-9 teratocarcinoma cells, with 50% inhibitory dose of 4.5 nM MTX, which makes it more active than free MTX (50% inhibitory dose of 15 nM). The drug-free antibody is not cytotoxic to F-9 cells at the concentrations used. The high efficacy of the conjugated drug may be due in part to the fact that anti-SSEA-1 antibody is an immunoglobulin M. MTX conjugated to nonspecific immunoglobulin M has little inhibitory effect (50% inhibitory dose of 150 nM). When acting on SSEA-1 negative cells, the two conjugates have only a small but identical effect. Thiamine pyrophosphate, an inhibitor of MTX transport, can prevent the cytotoxicity of the free MTX but not that of the anti-SSEA-1 conjugate. Leupeptin, an inhibitor of lysosomal protease, can partially protect F-9 cells against the antibody conjugate but not against free MTX. These results indicate that the MTX antibody conjugate binds specifically to F-9 cells, and is internalized and intracellularly degraded to release a small molecular active drug. Pretreatments of F-9 cells for 1 h with unlabeled antibody inhibits the subsequent uptake of identical concentration of labeled conjugate. The rate of internalization, however, regains almost normal values within 4 h, indicating a rapid reappearance of free antigenic sites at the cell surface.

Animals↗
PubMed1985

A method of membrane preparation for immunoassay.

Membranes prepared from a variety of solid tissues were used as solid-phase antigens for ELISA or RIA after fixation onto polylysine-primed 96-well plates. The preservation of antigens in these membrane preparations was tested by reactivity in ELISA using 2 monoclonal antibodies: W6/32, which recognizes an HLA framework antigen (a protein antigen) and anti-SSEA-1, directed to a carbohydrate antigen carried on glycoproteins. Levels of antigen deposition and usefulness as solid-phase antigens were assessed for ELISA as compared to RIA. Coated plates may be frozen for many months with preservation of antigenic activity. This method is relatively simple, rapid, and is useful for preparation of tissue antigens for immunoassay, especially for screening monoclonal antibodies.

Animals↗
PubMed1985

Tumor location using F(ab')2 mu from a monoclonal IgM antibody: pharmacokinetics.

A monoclonal IgM antibody (anti-SSEA-1) and its divalent antigen-binding peptic fragment [F(ab')2 mu] were compared as in vivo tumor localization reagents in mouse teratocarcinomas. F(ab')2 mu is cleared more rapidly than whole antibody from the whole body, blood, and all tested organs (t1/2 for whole antibody approximately 18 hr; t1/2 for F(ab')2 mu, 12 hr). A corresponding average improvement in tumor-to-tissue ratio is observed 48 hr after injection and earlier. However, the affinity of the F(ab')2 mu for antigen is much lower, and a smaller fraction of the antibody fragment is retained in the tumor than with whole antibody. The fragment was not retained by animals bearing nonantigenic tumors.

Animals↗
PubMed1984

Tumor radioimmunolocation: differential antibody retention by antigenic normal tissue and tumor.

On the assumption that a high specificity would be required for in vivo tumor location and drug targeting, much labor has been expended to produce monoclonal antibodies directed to antigens found primarily on tumors. However, for at least one tumor-associated antigen, stage-specific embryonic antigen 1 (SSEA-1), the concentration of the target antigen need not be higher in the tumor than that found in normal tissues for successful use of the corresponding monoclonal antibody in in vivo tumor location; rather, accessibility and antibody metabolism are more important.

Animals↗
PubMed1976

HLA membrane antigens: sequencing by intrinsic radioactivity.

Radiochemical sequence data are presented for the amino termini of mixed HLA antigens A1 and B8 isolated from a human lymphoblastoid cell line. Cells were labeled intrinsically in small-scale tissue culture with 14C-labeled amino acids and [14C]pyruvate. The specific activities obtained were sufficiently high and uniform to permit direct radiochemical sequencing of the antigens. HLA antigens were isolated by adsorption to a solid-phase anti-beta2-microglobulin immunoadsorbent followed by electrophoresis. The method should be generally applicable and useful for sequencing all parts of the molecule.

Amino Acid Sequence↗
PubMed1974

Chloral hydrate: a solvent for biological membranes.

A buffer system conitainin chloral hydrate, taurine, and bromopyridinium lactate was used to dissolve several biological membranes and separate their protein components by polyacrylamide gel electrophoresis. This solvent system was capable of separating molecules of similar size on the hasis of their charge and allows easy recovery of the proteins Thus. aqueous chloral hydrate is an effective solvent for biological membranes.

Animals↗