Use of Chlorotoxin for Targeting of Primary Brain Tumors1

Gliomas are primary brain tumors that arise from differentiated glial cells through a poorly understood malignant transformation. Although glioma cells retain some genetic and antigenic features common to glial cells, they show a remarkable degree of antigenic heterogeneity and variable mutations in their genome. Glioma cells have recently been shown to express a glioma-specific chloride ion channel (GCC) that is sensitive to chlorotoxin (CTX), a small peptide purified from Leiurus quinquestriatus scorpion venom [N. Ullrich et al.. Neuroreport, 7: 10201024, 1996; and N. Ullrich and H. Sontheimer, Am. J. Physiol. (Cell Physiol.), 270: C1511-C1521, 1996]. Using native and recombinant 125Ilabeled CTX, we show that toxin binding to glioma cells is specific and involves high affinity [dissociation constant (A',,i = 4.2 nM] and low affinity (Ka = 660 nM) binding sites. In radioreceptor assays, I25l-labeled CTX binds to a protein with M, = 72,000, presumably GCC or a receptor that modulates GCC activity. In vivo targeting and biodistribution experiments were obtained using I25Iand I3ll-labeled CTX injected into severe com bined immunodeficient mice bearing xenografted gliomas. CTX selectively accumulated in the brain of tumor-bearing mice with calculated brain: muscle ratios of 36.4% of injected dose/g (ID/g), as compared to 12.4%ID/g in control animals. In the tumor-bearing severe combined immunodeficient mice, the vast majority of the brain-associated radioac tivity was localized within the tumor (tumor: muscle ratio, 39.13% ID/g; contralateral braimmuscle ratio, 6.68%ID/g). Moreover, 131I-labeled CTX distribution, visualized through in vivo imaging by gamma ray camera scans, demonstrates specific and persistent intratumoral localization of the radioactive ligand. Immunohistochemical studies using biotinylated and fluorescently tagged CTX show highly selective staining of glioma cells in vitro, in situ, and in sections of patient biopsies. Comparison tissues including normal human brain, kidney, and colon were consistently negative for CTX immunostaining. These data suggest that CTX and CTX-conjugated mol ecules may serve as glioma-specific markers with diagnostic and thera peutic potential.