Alexander Bruskin

Cellular processing of 125I- and 111in-labeled epidermal growth factor (EGF) bound to cultured A431 tumor cells

Low molecular weight of epidermal growth factor (EGF) enables better intratumoral penetration in comparison with larger targeting proteins, but the cellular retention of EGF-associated radioactivity is poor for directly iodinated EGF. An attempt was made to improve intracellular retention by the use of metal-diethylenetriaminepentaacetic acid or nonphenolic linker (N-succinimidyl-para-iodobenzoate) as labeling agents. The use of nonphenolic linker did not improve retention of the radioactivity in A431 carcinoma cell line. The use of the radiometal label provided an appreciable prolongation of radioactivity residence inside the cell.

Radiobromination of closo-dodecaborate anion. Aspects of labelling chemistry in aqueous solution using Chloramine-T

Summary Closo-dodecaborate dianion is a three-dimensional aromatic inorganic molecule, which can be easily halogenated forming a stable halogen-boron bond. Derivatives of closo-dodecaborate were considered as a convenient chemical form of delivery of enriched 10B to malignant tumors for boron neutron capture therapy (BNCT). Some properties of closo-dodecaborate (hydrophilicity, strength of halogen-boron bond, charge at lysosomal pH) make it attractive as a potential prosthetic group for attachment of radioactive halogens to tumor-targeting proteins. Bromine radioisotopes possess a variety of useful nuclear characteristics, and can be used in different areas of clinical diagnostics and therapy. In this work, a basic chemistry of closo-dodecaborate radiobromination was studied. It was found, that di(triethylamonium) dodecahydro-closo-dodecaborate can be labelled in high yield, more then 90%, in a wide pH range. By decreasing the pH, the bromination can be directed to closo-dodecaborate in the presence of phenolic compounds. The results of the study indicate a possibility of using the radioactive bromine label for investigation of pharmacokinetics of boronated compounds for BNCT.

[111In]Bz-DTPA-hEGF : Preparation and in vitro characterization of a potential anti-glioblastoma targeting agent

The overexpression of epidermal growth factor receptors, EGFR, in glioblastomas is well documented. Hence, the EGFR can be used as target structure for a specific targeting of glioblastomas. Both radiolabeled anti-EGFR antibodies and the natural ligand EGF are candidate agents for targeting. However, EGF, which has a rather low molecular weight (6 kDa), might have better tissue penetration properties through both normal tissue and tumors in comparison with anti-EGF antibodies and their fragments. The aim of this study was to prepare and evaluate in vitro an EGF-based antiglioma conjugate with residualizing label. Human recombinant EGF (hEGF) was coupled to isothiocyanate-benzyl-DTPA. The conjugate was purified from unreacted chelator using solid-phase extraction and labeled with (111)In. The labeling yield was 87% +/- 7%. The label was reasonably stable; the transchelation of (111)In to serum proteins was about 5% after incubation at 37 degrees C during 24 hours. The obtained [(111)In]benzyl-DTPA-hEGF conjugate was characterized in vitro using the EGFR expressing glioma cell line U343MGaCl2:6. The binding affinity, internalization, and retention of the conjugate were studied. The conjugate had receptor specific binding and the radioactivity was quickly internalized. The intracellular retention of radioactivity after interrupted incubation with conjugate was 71% +/- 1% and 59% +/- 1.5% at 24 and 45 hours, respectively. The dissociation constant was estimated to 2.0 nM. The results indicate that [(111)In]benzyl-DTPA-hEGF is a potential candidate for targeting glioblastoma cells, possibly using locoregional injection.

[177Lu]Bz-DTPA-EGF: Preclinical characterization of a potential radionuclide targeting agent against glioma.

Patients with glioblastoma multiforme have a poor prognosis due to recurrences originating from spread cells. The use of radionuclide targeting might increase the chance of inactivating single tumor cells with minimal damage to surrounding healthy tissue. As a target, overexpressed epidermal growth factor receptors (EGFR) may be used. A natural ligand to EGFR, the epidermal growth factor (EGF) is an attractive targeting agent due to its low molecular weight (6 kDa) and high affinity for EGFR. 177Lu (T(1/2) = 6.7 days) is a radionuclide well suited for treatment of small tumor cell clusters, since it emits relatively low-energy beta particles. The goal of this study was to prepare and preclinically evaluate both in vitro and in vivo the [177Lu]Bz-DTPA-EGF conjugate. The conjugate was characterized in vitro for its cell-binding properties, and in vivo for its pharmacokinetics and ability to target EGFR. [177Lu]Bz-DTPA-EGF bound to cultured U343 glioblastoma cells with an affinity of 1.9 nM. Interaction with EGFR led to rapid internalization, and more than 70% of the cell-associated radioactivity was internalized after 30 minutes of incubation. The retention of radioactivity was good, with more than 65% of the 177Lu still cell-associated after 2 days. Biodistribution studies of i.v. injected [177Lu]Bz-DTPA-EGF in NMRI mice demonstrated a rapid blood clearance. Most of the radioactivity was found in the liver and kidneys. The liver uptake was receptor-mediated, since it could be significantly reduced by preinjection of unlabeled EGF. In conclusion, [177Lu]Bz-DTPA-EGF seems to be a promising candidate for locoregional treatment of glioblastoma due to its high binding affinity, low molecular weight, and ability to target EGFR in vivo.