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Biomedical subjects

M F Hawthorne

Publications and source records attributed to M F Hawthorne.

34 records · Page 2Linked to original sources
PubMed1995

Preparation and properties of nido-carborane-specific monoclonal antibodies for potential use in boron neutron capture therapy for cancer.

As the first step of a research program aimed at developing a bispecific monoclonal antibody system for the delivery of boron-rich molecules to tumor cells for boron neutron capture therapy, monoclonal antibodies (mAbs) were produced against an anionic nido-carborane derivative, 4-[7,8-dicarbadodecahydroundecaborat(-1)-7-yl]butanoic acid. Two IgG subclass mAbs, designated HAW101 and HAW102, were identified that specifically bound the anionic nido-carborane hapten, as well as a variety of other anionic nido-carborane cage derivatives. By using surface plasmon resonance technology, the affinity constants of HAW101 and HAW102 were determined to be 1.9 x 10(9) and 6.8 x 10(8) M-1, respectively. A diverse array of 7-substituted and 7,8-disubstituted anionic nido-carborane derivatives reacted with the mAb HAW101 in competition ELISA, whereas anionic closo-polyhedral boranes showed negligible binding, suggesting a role for the open nido-carborane cage structure. These results suggest that mAbs such as HAW101, which bind anionic nido-carboranes, are useful in the development of bispecific mAbs for specific targeting and enhanced boron delivery to tumor sites.

Antibodies, Monoclonal↗
PubMed1995

Selective boron delivery to murine tumors by lipophilic species incorporated in the membranes of unilamellar liposomes.

The nido-carborane species K[nido-7-CH3(CH2)15-7,8-C2B9H11] has been synthesized for use as an addend for the bilayer membrane of liposomes. Small unilamellar vesicles, composed of distearoylphosphatidylcholine/cholesterol, 1:1, and incorporating K[nido-7-CH3(CH2)15-7,8-C2B9H11] in the bilayer, have been investigated in vivo. The time-course biodistribution of boron delivered by these liposomes was determined by inductively coupled plasma-atomic emission spectroscopy analyses after the injection of liposomal suspensions in BALB/c mice bearing EMT6 mammary adenocarcinomas. At the low injected doses normally used (approximately 5-10 mg of boron per kg of body weight), peak tumor boron concentrations of approximately 35 micrograms of boron per g of tissue and tumor/blood boron ratios of approximately 8 were achieved. These values are sufficiently high for the successful application of boron neutron capture therapy. The bilayer-embedded boron compound may provide the sole boron source or, alternatively, a concentrated aqueous solution of a hydrophilic boron compound may also be encapsulated within the liposomes to provide a dose enhancement. Thus, the incorporation of both K[nido-7-CH3(CH2)15-7,8-C2B9H11] and the hydrophilic species, Na3[1-(2'-B10H9)-2-NH3B10H8], within the same liposomes demonstrated significantly enhanced biodistribution characteristics, exemplified by maximum tumor boron concentrations of approximately 50 micrograms of boron per g of tissue and tumor/blood boron ratios of approximately 6.

Animals↗
PubMed1994

Na3[B20H17NH3]: synthesis and liposomal delivery to murine tumors.

The polyhedral borane ion [n-B20H18]2- reacts with liquid ammonia in the presence of a suitable base to produce an apical-equatorial (ae) isomer of the [B20H17NH3]3- ion, [1-(2'-B10H9)-2-NH3B10H8]3-. The structure of this product has been confirmed by 11B NMR spectroscopy and x-ray crystallography. This species undergoes acid-catalyzed rearrangement to an apical-apical (a2) isomer, [1-(1'-B10H9)-2-NH3B10H8]3-, whose structure has been determined by 11B NMR spectroscopy. The sodium salts of both the ae and the a2 isomers of [B20H17NH3]3- have been encapsulated within small unilamellar liposomes, composed of distearoyl phosphatidylcholine/cholesterol (1:1), and investigated as boron-delivery agents for boron neutron capture therapy (BNCT) of cancer. The biodistribution of boron was determined after the injection of liposomal suspensions into BALB/c mice bearing EMT6 tumors. Both [B20H17NH3]3- isomers exhibited excellent tumor uptake and selectivity at very low injected doses, achieving peak tumor boron concentrations of 30-40 micrograms of B/g of tissue and tumor/blood boron ratios of approximately 5. The enhanced retention of the [B20H17NH3]3- isomers by EMT6 tumors may be attributed to their facile intracellular oxidation to an extremely reactive NH3-substituted [n-B20H18]2- ion, the electrophilic [B20H17NH3]- ion. Both isomers of [B20H17NH3]3- are at least 0.5 V more easily oxidized than other previously investigated species containing 20 boron atoms. In another experiment, [ae-B20H17NH3]3- was encapsulated in liposomes prepared with 5% PEG-2000-distearoyl phosphatidylethanolamine in the liposome membrane. As expected, these liposomes exhibited a longer circulation lifetime in the biodistribution experiment, resulting in the continued accumulation of boron in the tumor over the entire 48-hr experiment and reaching a maximum of 47 micrograms of B/g of tumor.

Animals↗
PubMed1993

Pharmacokinetics of an anti-carcinoembryonic antigen monoclonal antibody conjugated to a bifunctional transition metal carborane complex (venus flytrap cluster) in tumor-bearing mice.

An anticarcinoembryonic antigen (CEA) monoclonal antibody, T84.66, has been conjugated to a metallocarborane complex (Venus flytrap cluster, VFC) containing 57Co. This radioimmunoconjugate, 57Co-VFC-T84.66, retained > 90% immunoreactivity, was stable in serum (7 days) and demonstrated good localization in LS174T tumor xenografts. Pharmacokinetics of 57Co-VFC-T84.66 in tumor-bearing mice were compared to T84.66 Mab conjugated with either DTPA or its benzylisothiocyanate derivative (BzDTPA) labeled with 111In. Whole-body half-life for VFC-T84.66 was less (t1/2 = 62 hr) than that for either DTPA-T84.66 (t1/2 = 157 hr) or BzDTPA-T84.66 (t1/2 = 167 hr). Blood clearance was similar for all three radioimmunoconjugates (t1/2 = 22 hr). Hepatic uptake of the radiolabel was rapid and remained constant for 7 days for both DTPA radioimmunoconjugates (DTPA radioimmunoconjugate = 13.7 +/- 1.5 %ID/g; BzDTPA radioimmunoconjugate = 10.4 +/- 1.7%ID/g). For VFC, however, liver radioactivity decreased from 19.1 +/- 0.6%ID/g at 1 hr to 0.9 +/- 0.1 %ID/g 7 days postinjection, suggesting a possible role for VFC radioimmunoconjugate in the imaging and therapy of liver metastases.

Animals↗
PubMed1992

Model studies directed toward the boron neutron-capture therapy of cancer: boron delivery to murine tumors with liposomes.

The successful treatment of cancer by boron neutron-capture therapy (BNCT) requires the selective concentration of boron-10 within malignant tumors. The potential of liposomes to deliver boron-rich compounds to tumors has been assessed by the examination of the biodistribution of boron delivered by liposomes in tumor-bearing mice. Small unilamellar vesicles with mean diameters of 70 nm or less, composed of a pure synthetic phospholipid (distearoyl phosphatidylcholine) and cholesterol, have been found to stably encapsulate high concentrations of water-soluble ionic boron compounds. The hydrolytically stable borane anions B10H10(2-), B12H11SH2-, B20H17OH4-, B20H19(3-), and the normal form and photoisomer of B20H18(2-) were encapsulated in liposomes as their soluble sodium salts. The tissue concentration of boron in tumor-bearing mice was measured at several time points over 48 h after i.v. injection of emulsions of liposomes containing the borane anions. Although the boron compounds used do not exhibit an affinity for tumors and are normally rapidly cleared from the body, liposomes were observed to selectively deliver the borane anions to tumors. The highest tumor concentrations achieved reached the therapeutic range (greater than 15 micrograms of boron per g of tumor) while maintaining high tumor-boron/blood-boron ratios (greater than 3). The most favorable results were obtained with the two isomers of B20H18(2-). These boron compounds have the capability to react with intracellular components after they have been deposited within tumor cells by the liposome, thereby preventing the borane ion from being released into blood.

Animals↗
PubMed1991

A transition metal complex (Venus flytrap cluster) for radioimmunodetection and radioimmunotherapy.

A novel transition metal complex, Venus flytrap cluster (VFC), is described for the preparation of radio-labeled antibodies. VFC contained 57Co, which was held tightly between the faces of two covalently bridged carborane ligands by cluster bonding of the metal with appropriate ligand orbitals. Anti-carcinoembryonic antigen monoclonal antibody T84.66 was conjugated to 57Co-VFC with full retention of immunological activity. Biodistribution studies in nude mice bearing carcinoembryonic antigen-producing tumors showed excellent tumor localization of 57Co-VFC-T84.66. The accumulation of radionuclide in normal liver was low and independent of dose, which may reflect the stability of the radionuclide complex. These results presage the use of VFC systems for binding transition metals that are clinically useful for radioimmunodiagnosis and radioimmunotherapy.

Animals↗
PubMed1984

Neutron-capture therapy of human cancer: in vivo results on tumor localization of boron-10-labeled antibodies to carcinoembryonic antigen in the GW-39 tumor model system.

Antibody against carcinoembryonic antigen (CEA) was conjugated with p-[1,2-dicarba-closo-[1-3H]do-decaboran(12)-2-yl] benzenediazonium ion by an azo-coupling reaction, resulting in 30 boron atoms per IgG molecule with no loss of antibody protein. Antibody immunoreactivity was not appreciably affected by this conjugation and was stable after incubation in vitro in hamster plasma for 24 hr. The efficacy of the boron-conjugated anti-CEA IgG for localizing selectively in CEA-containing human colonic carcinomas propagated in the hind leg musculature of hamsters was evaluated by labeling the antibodies with 131I and determining distribution of the radioactivity in vivo. The results show that the boron-conjugated antibodies retain selective localization in the tumors, thus indicating their suitability for transporting boron-10 to tumors for use in neutron-capture therapy of cancer.

Antibodies, Neoplasm↗
PubMed1982

Neutron-capture therapy of human cancer: in vitro results on the preparation of boron-labeled antibodies to carcinoembryonic antigen.

Two samples of 2-phenyl-1,2-dicarba-closo-[1-3H]dodecaborane(12) were prepared by treating 1-lithio-2-phenyl-1,2-dicarba-closo-dodecaborane(12) with 3H2O (0.1 and 5.0 Ci/ml, respectively). These tritiated phenylcarborane samples were subsequently converted to corresponding samples of p-[1,2-dicarba-closo-[1-3H]dodecaboran(12)-2-yl]benzenediazonium ion ([3H]DBD) suitable for azo-coupling reactions. Reaction of the two tritiated diazonium ion samples with 2-napthol resulted in the formation of an azo dye (epsilon = 1.98 X 10(4) M-1 cm-1 at 485 nm). Experiments relating absorbance to 3H activity proved the two [3H]DBD sources to have 3.81 X 10(11) and 2.45 X 10(13) cpm of 3H per mol of tritiated carborane substituent. Purified antibodies to carcinoembryonic antigen were coupled to the [3H]DBD and, after extensive dialysis, the average number of carborane moieties per antibody molecule was determined by measuring the 3H activity associated with a known protein concentration. Further examination of these tritiated carborane-labeled antibodies by affinity chromatography proved that boron labeling did not destroy their immunoreactivity. Correlations of azo-coupling conditions (reactant ratios, pH) with immunoreactivity and antibody protein recovery are presented.

Antibodies, Neoplasm↗
PubMedDate not supplied

Conjugation of phenyl isothiocyanate derivatives of carborane to antitumor antibody and in vivo localization of conjugates in nude mice.

A systematic study of the conjugation of 1-(p-isothiocyanatophenyl)-1,2-dicarba-closo-[2-3H]dodeca borane(12), 3H-1, and 7-(p-isothiocyanatophenyl)dodecahydro-7,8-dicarba-nido -[8-3H] undecaborate(1-)ion, 3H-2, to the murine monoclonal anti-CSAp antibody, Mu-9, was carried out to compare charged and uncharged boron cages in their effect upon antibody loading. Approximately one neutral cage and four of the anionic cages were successfully linked to antibody in two separate conjugates which were subsequently radioiodinated and evaluated in vivo. No significant loss of the antibody or its immunoreactivity was observed in either case. In nude mice bearing GW-39 tumor xenografts the conjugate containing the anionic carborane derivative showed a reduced tumor uptake although the tumor:non-tumor ratio was similar to that of the native antibody. The carborane cage in 2 was radiolabeled with 125I followed by attempts to purify and conjugate product 4 to a model goat IgG protein. This exploratory conjugation study was undertaken as a prelude to linking new conjugation reagents, which contain multiple anionic boron cages, to antitumor antibodies. The latter conjugates are required to maximize boron loading for the purpose of neutron-capture therapy.

Animals↗
PubMedDate not supplied

Novel carboranyl amino acids and peptides: reagents for antibody modification and subsequent neutron-capture studies.

A new alpha-amino acid derivative incorporating the 1,2-dicarba-closo- dodecarborane(12) cage, namely 5-(2-methyl-1,2-dicarba-closo-dodecarborane(12)-1-yl)- 2-aminopentanoic acid (2), was synthesized by the alkylation of the benzophenone Schiff's base of glycine methyl ester with 3-(2-methyl-1,2-dicarba-closo-dodecaborane(12)-1-yl)pr opyl iodide (8). This amino acid was employed in the synthesis of peptide derivatives such as 19-21 using solid-phase Merrifield methods. Dipeptide 19 was converted to a water-soluble ionic derivative by the pyrrolidine-mediated carborane cage degradation reaction followed by cation exchange to afford sodium salt 22. Dansylation of 22 with dansyl chloride yielded fluorescence-labeled dipeptide 23. Undecapeptide 21 was dansylated while still anchored to the Merrifield resin. Following its cleavage from the resin with hydrogen fluoride, product 25 was acetylated to block the free amino group on the lysine residue and then converted to water-soluble derivative 27. Trial conjugations of dipeptide 23 and undecapeptide 27 to T84.66, an anti-CEA antibody, were carried out by means of carboxyl activation with N-hydroxysulfosuccinimide and N,N-diisopropylcarbodiimide. Studies of the chemical syntheses of these and other peptide derivatives and the conjugation of 23 and 27 to the antibody are described.

Amino Acids↗
PubMedDate not supplied

Carboranyl peptide-antibody conjugates for neutron-capture therapy: preparation, characterization, and in vivo evaluation.

Two model peptides rich in boron and prepared by Merrifield syntheses, dansyl.(nido-CB)2, (1) and dansyl.(nido-CB)10.Lys.Ac (2), where nido-CB represents the alpha-amino acid [nido-7-CH3-8-(CH2)3CH-(NH2)COOH-7,8-C2B9H10]-, were conjugated with the anti-CEA mAb T84.66 using peptide active ester reagents. The dansyl groups provided a means of fluorimetric analysis of mAb conjugates which was augmented by conventional amino acid analyses for nido-CB. The conjugate of 1 contained an average of 63 B atoms per mAb molecule. The mAb conjugate of 2 was chromatographically separated into a strongly fluorescent high molecular weight aggregated fraction (HMW) and a less intensely fluorescent monomeric fraction. Both fractions retained immunoreactivity. The HMW species contained an average of ca. 490 B atoms/mAb molecule, as determined by amino acid analysis. Biodistribution data were collected using nude mice bearing LS174T xenografts and 125I-labeled mAb conjugates. While the lightly B-loaded dipeptide conjugate gave biodistribution results which resembled those of native T84.66 mAb, the undecapeptide conjugate displayed greatly enhanced liver uptake and decreased tumor accretion. These results suggest that as the boron-containing burden on the supporting immunoprotein is greatly increased, as in the case of the T84.66-2 conjugate, loss of circulating conjugate to liver effectively competes with the desired tumor localization. Means which might be taken to circumvent this difficulty have been described elsewhere (ref 15).

Animals↗
PubMedDate not supplied

Synthesis and characterization of oligomeric nido-carboranyl phosphate diester conjugates to antibody and antibody fragments for potential use in boron neutron capture therapy of solid tumors.

Antibodies conjugated to oligomeric carboranyl compounds have a high potential as target species for boron neutron capture therapy (BNCT) of solid tumors. As a first step toward developing conjugates with BNCT capabilities, an oligomeric nido-carboranyl phosphate diester (Kane, R. R., Dreschel, K., and Hawthorne, M.F. (1993) J. Am. Chem. Soc. 115, 8853-8854), CB10 (10 nido-carboranes containing 90 boron atoms) with a pseudo-5'-terminal amino group, was conjugated to the anticarcinoembryonic antigen antibody T84.66 and its F(ab') fragment. The homobifunctional linker disuccinimidyl suberate (DSS) was coupled to CB10 via its 5'-terminal amino group followed by removal of excess linker with organic solvent extraction and conjugation with intact antibody. Similarly, the heterobifunctional linker, m-maleimidobenzoyl-N-hydroxysuccinimide (MBS), was coupled to CB10 and conjugated to the hinge region sulfhydryl of the F(ab') fragment of T84.66. The extent of reaction was monitored by the mobility shift of CB10-antibody conjugate on native polyacrylamide gels and the increased susceptibility of the CB10-antibody conjugate to staining with silver nitrate. CB10 was also labeled with radioiodine (131I) in a solid phase reaction with iodogen and used in double-label studies with 125I-labeled antibody. Although free CB10 bound very tightly to gel filtration media such as Sephadex G-25, the CB10-antibody conjugate passed through freely. After separation of CB10-antibody conjugate from free CB10 on Sephadex G-25, molar incorporations of CB10 were calculated. At a molar ratio of 10:1 (CB10:T84.66), greater than 90% of T84.66 and 30% of its F(ab)' fragment were conjugated to CB10.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMedDate not supplied

Bispecific antibody mediated targeting of nido-carboranes to human colon carcinoma cells.

Boron neutron capture therapy, a binary form of cancer treatment, has the potential to deliver potent cytotoxic radiation to tumor cells with minimal collateral damage to normal tissues if methods for the selective accretion of elevated concentrations of boron-10 in tumor can be developed. In this regard, a monoclonal antibody with dual specificity, for both anionic boron cluster compounds (nido-carboranes) and a tumor-associated antigen (carcinoembryonic antigen, CEA), was produced. The specific binding of a nido-carborane to CEA-expressing tumor cells was achieved using this bispecific antibody. The ability of this bispecific antibody to concentrate selectively at tumor sites in vivo has also been demonstrated, thus suggesting its potential for sequestering boron-rich compounds in tumors.

Antibodies, Bispecific↗